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ILLINOIS BIOLOGICAL 
MONOGRAPHS 


Vol. XI July, 1927 No. 3 


EDITORIAL COMMITTEE 


STEPHEN ALFRED FORBES Homer Lr Roy SHANTZ 


HENRY BALDWIN WARD 


PUBLISHED UNDER THE 
AUSPICES OF THE GRADUATE SCHOOL BY 
THE UNIVERSITY OF ILLINOIS 


CopyricHtT, 1928, By THE UNIVERSITY OF ILLINOIS 
DISTRIBUTED NOVEMBER 17, 1928 


THE STRUCTURE AND DEVELOPMENT 
OF CORALLOBOTHRIUM 


With Descriptions of Two New Fish Tapeworms 


WITH FIVE PLATES 


BY 
HIRAM ELI ESSEX 


Contributions from the 
Zoological Laboratory of the University of Illinois 
under the direction of Henry B, Ward 
No. 323 


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Tn trod UCtion sane meer pieces 5 ce aie csacry ieton agieyiva tea sacs fee ven ieeesge Gio eae 7 
MaterialsvandeWiethods so. cs 2 seroca.c deena tee Sad otre Hw wimawromnis cent ote no 8 
The'Genus Corallobothrivm. «ccc. ccccc cscs ecwieacas ees ws ce gseeaw se eedes geese? 9 
Corallobothrium giganteum NOV. SPEC... 2. eee ii 
Reproductive Organs). (6 cise. fe cure nctewsaiGlem secs bas eaten eG cee secede 16 
Amphitypy.2s2sc:-<s9ss-- Sein RE ON eee Dine aoe aioe eae 20 
Corallobothrium fimbriatum nov. spec......... Nite Mee wsinar cease Ree e Ss 21 
Reproductive Organs.................. Pe Fete rae eye 24 
Distribution, Abundance and Seasonal Occurrence.................00000000 000000 30 
Wiferbustory, of Corallobothrium’ 25. cvaiavaiiesani ea ayer teeta G.ewe sate stets ee werrtrenes 32 
Observations Concerning the Eggs... 0... eee 33 
SXperiments swith: thee eS 5.5 tang csinijensisss alae enesers ols eteye eps Orel ceeael@iepansleoe ce als 36 
The: First. Intermediate Host: «cen cua cece s cote Sted cre hee ee neon so 37 
Development of Corallobothrium fimbriatum in Cyclops.................. . 40 
Development of Corallobothrium giganteum in Cyclops.................. . 4 
The Second Intermediate Host............. 2... cece cece eect enn eee . 46 
(Phe Plerocercoid Larvae aa oa ¢acac; emit ein ete see wets du eaclee eS, er 49 
Comparison of Proteocephalus and Corallobothrium........................ ; fol 
Comparison with Diphyllobothrium latum......0.0. 0000 oe ee ogee os) 
Early Development in Bothriocephalids and Proteocephalids................ , 54 
Afhnities of the Proteocephalids: :..2.:3..0ccc0s0cceesceeewsendweeeeewecnebecuies 57 
Summary.......... ee ee oa lado ts tion Reese Re Tae er ee ee 58 
Bibliography.................... RE On ear oN Ao ee eee eee .. 60 


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261] STRUCTURE AND DEVELOPMENT OF CORALLOBOTH RIUM—ESSEX 7 


INTRODUCTION 


Representatives of the genus Corallobothrium from American fish 
have never been described. Marshall and Gilbert (1905) report two 
members of this genus taken from Ameiurus melas (black bullhead) 
caught in the lakes at Madison, Wisconsin. La Rue (1914) refers to an 
undescribed species of Corallobothrium encysted in the livers of A. melas 
and A. nebulosus (common bullhead) from the Illinois River. Ward 
(1918) noted the presence of a species of Corallobothrium in Ictalurus 
punctatus (channel-cat) at Milford, Nebraska. Aside from these inci- 
dental references, the American literature contains no information on 
this interesting group of fish tapeworms. 

The acquaintance of the writer with these cestodes began during 
the summer of 1925 while a survey of the parasites of fish from the rivers 
and lakes in the upper Mississippi and Missouri basin was in progress 
(Essex and Hunter, 1926). At that time cestodes belonging to Corallo- 
bothrium were found in the following hosts: Ameturus melas, Leptops 
olivaris (mud-cat), and Ictalurus punctatus. 

At the suggestion of Dr. Henry B. Ward an intensive study was begun 
in the spring of 1926 on the species of Corallobothrium infesting 7. punc- 
tatus. As the investigation progressed it came to assume three distinct 
phases: (1) taxonomy and morphology; (2) seasonal occurrence; (3) life- 
cycle. It was discovered that two new species of Corallobothrium were 
represented in my collections. These two species are described and named 
in the present paper. No work having been reported previously on the 
seasonal occurrence of any fish tapeworm in America, an effort was made 
to obtain all possible information on this phase of the problem. This 
study has shed some light on the biological relations between the para- 
sites and their hosts. Up to this time only a few experimental investiga- 
tions on the life-cycle of fish cestodes have been reported by European 
workers and none whatever by workers in America. Since such studies 
are of especial biological significance, and since complete information 
on the developmental history of fish parasites is of great assistance to 
fish culturists in their efforts to combat parasitic diseases among fish, 
all data that I have secured on this phase of the investigation are pre- 
sented in this paper. 

My most sincere thanks are extended to Professor Ward for his in- 
spiration, guidance and helpful criticism in this work. I am deeply grate- 
ful for the use of the zoological laboratories of Rockford College which 
were made available to me, during the summer of 1926, through the 


8 ILLINOIS BIOLOGICAL MONOGRAPHS [262 


kindness of Dr. Ruth Marshall and the generosity of the college authorities. 
I also acknowledge here the invaluable assistance rendered by Dr. David 
H. Thompson of the Illinois Natural History Survey, who furnished me 
several shipments of catfish for examination. Thanks are also due Mr. 
R. E. Richardson for the identification of minnows and for a collection 
of parasites from Ictalurus punctatus, and to Dr. S. A. Forbes for the 
use of unpublished data on the food of I. punctatus collected by the Illi- 
nois Natural History Survey. 


MATERIAL AND METHODS 


The parasites were killed in corrosive sublimate, Bouin’s solution and 
4 per cent formol. The first two gave good results, but specimens killed 
in formol were difficult to stain. Ehrlich’s or Delafield’s hematoxylin 
was used for totos and sections. Counterstaining was done with eosin 
or orange G. 

Collection of the adult cestodes offers no difficulties, but it is a long 
and tedious struggle to find the smallest larval forms. A method was 
developed whereby plerocercoids which measured only 0.25 mm long 
could be collected with a saving of time and effort. Briefly stated, the 
procedure is as follows: (1) large parasites and particles of debris 
are removed; (2) remainder of intestinal contents scraped into quart 
jar of water and shaken thoroughly; (3) the mass is strained through a 
small plankton net; (4) the plankton net placed under faucet, running 
water allowed to play on contents until all possible material is washed 
through the net; (5) the residue is turned into a watchglass and examined 
under a binocular. This method is equally successful in collecting small 
trematodes. 

A rapid method of reconstruction was developed in connection with 
the work on these cestodes. A plate of glass one foot square was placed 
under the camera lucida. With a wax pencil the structure to be recon- 
structed from a given section was traced on the glass. With plasticene 
the tracings were modeled. The next section was then oriented accord- 
ing to guide lines and the required tracings made from it. By means 
of blocks of plasticene the structures modeled from the second, third, 
and later sections were placed according to scale above those previously 
modeled. When completed, the organs were represented in three di- 
mensions and in their proper relation. This method was employed in 
making the models represented by figures 39 and 41. 


263] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 9 


THE GENUS CORALLOBOTHRIUM 


The genus Corallobothrium was created by Fritsch in 1886 to ac- 
commodate a species of cestode from Malapterurus electricus, an elec- 
tric catfish of Egypt. Because of the resemblance of the scolex to the 
structure of an Oculina-like coral, he named the parasite Corallobothrium 
and called the species C. solidum. It was his opinion that this genus 
represented a connecting link between the Bothriocephalan and Taenian 
cestodes. He gave the following diagnosis for the genus: ‘“‘Caput bothrio 
uno terminali, fere plano, ovali lateribus attenuatis, superficie et margine 
crispo. Acetabula quattuor cruciatim posita, in bothrii medio profunde 
inserta. Collum nullum. Corpus articulatum, depressum, subaequale 
vel retrorsum angustatum. Organa genitalia typica, orificia marginalia 
involuta.’’ Fritsch says that the form of the scolex recalls that of Caryo- 
phyllaeus, ‘‘nur ist die Sauggrube in ihrer ovalen nach beiden Seiten 
leicht vershmalerten Gestalt, viel regelmassiger gebildet’’; that the ace- 
tabular suckers are entirely obscured by the folds of the scolex and are 
revealed only in sections. The suckers agree in structure and arrange- 
ment with those of Taenia; the character of the whole body is strikingly 
solid and strong. 

No other species was referred to the genus Corallobothrium before 
the work of Riggenbach (1896), who described a cestode from Pime- 
lodus pati, a siluroid of Paraguay. This parasite was designated as Corallo- 
bothrium lobosum. Fuhrmann (1916) redescribed this form and gave a 
short redescription of C. solidum. On the basis of the cortical arrange- 
ment of the testes and vitellaria he removed C. Jobosum from the genus 
Corallobothrium and created for it a new genus, namely, Rudolphiella. 
Later Fuhrmann and Baer (1925) reduced Rudolphiella to synonomy, 
and placed R. Jobosa in the genus Ephedrocephalus. 

Braun (1895) accepts the genus Corallobothrium and to Fritsch’s 
diagnosis adds that neither hooks nor spines are present; that excretory 
vessels have secondary openings in the proglottids; and that the hosts 
are tropical or subtropical bony-fish. La Rue (1914) adds to Braun’s 
diagnosis that the folds and lappets of the scolex may enclose the suckers 
as in a corolla and that no rostellum is present. He omits the secondary 
openings of the excretory system, given by Braun, and gives the habitat 
as the intestinal tract of the Siluridae. 

The validity of the genus Corallobothrium was never questioned 
until the appearance of two papers by Woodland (1925, 1925a) in which 
he proposes to delete such genera as Corallobothrium, Choanoscolex, 


10 ILLINOIS BIOLOGICAL MONOGRAPHS [264 


Acanthotaenia and Gangesia and make them synonyms of Proteocepha- 
lus because they were founded on scolex characteristics. This author 
regards such characters as of specific value only, and proposes that the 
various arrangements of the reproductive organs with reference to the 
inner longitudinal muscle sheath shall constitute the bases for the desig- 
nation of genera. In Woodland’s opinion the scolex characters are of 
secondary importance as compared to the structure of the proglottids. 
However, if the early development of the cestodes is accepted as a guide 
in determining the order of sequence which is followed in arriving at the 
adult organization, and if those structures which appear first in the develop- 
ment of the individual are considered as having arisen first in the phylo- 
geny of the group, then the scolex must be acknowledged as one of the 
most fundamental structures of the adult organism. In the development 
of Taenia and related groups, the scolex and a more or less indifferent 
neck are the only adult structures found in the cysticercus, and from them 
arise allotheradult organs. In the development of the two new species des- 
cribed in this paper the scolex is likewise the first adult structure to ap- 
pear, being differentiated at the end of 10 to 12 days development. When 
considered from this point of view, there is certainly sufficient reason for 
attaching generic value to the character of the scolex. Fuhrmann (1916) 
doubtless recognized this when he created a separate genus for Goezeella 
siluri instead of referring it to the genus Monticellia; the only essential 
difference between the two genera being that the former possesses a 
scolex of the Corallobothrium type. Furthermore, the scolex has long 
been used by helminthologists as a generic character, and to discard it 
now would result in more confusion of the nomenclature than seems to 
be justified. A recent publication by Stiles and Hassell (1926) contains 
the following statement: ‘‘The genus Taenia, in its modern concept, 
can be subdivided into at least three groups to which either subgeneric 
or generic rank may be given (according to one’s personal point of view 
in respect to generic values).”’ A key follows in which the genera Taenia, 
Hydatigera and Taeniarhynchus are separated entirely on the basis of 
the scolex while Taenia and Taeniarhynchus are separated on the nature 
of the rostellum alone. 

Since the characters which constitute a genus are considered largely 
a matter of personal viewpoint, Woodland is hardly justified in deleting 
genera which have been accepted previously by the leading workers in 
helminthology because, from his point of view, he does not regard the 
characters on which those genera were founded as of generic value. There- 
fore, I have followed Braun, Fuhrmann and others in accepting the 
genus Corallobothrium and the two new species described in this paper 
have been referred to that genus. 


265] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 11 


Corallobothrium is closely related to the genus Proteocephalus. The 
internal organization of the two genera is very similar. The scolex, how- 
ever, is so distinctive that species belonging to the two genera can be 
separated by an examination of the scolex alone. Another character 
by which the species of Corallobothrium, which I have studied, may be 
separated from those of Proteocephalus, is in the position of the vagina 
with reference to the cirrus-pouch. In Corallobothrium giganteum and C. 
jimbriatum, the vagina in successive segments varies and may be either 
anterior or posterior to the cirrus-pouch. The descriptions of C. solidum 
are silent on this point, but if this condition is true of it also, there exist 
two distinct points of difference, scolex and position of vagina, between 
the species of Corallobothrium and Proteocephalus. Regarding the latter 
La Rue (1914) makes the following statement: ‘‘In Proteocephalus the 
vagina is usually anterior to the cirrus-pouch although there are a few 
species in which it is regularly posterior.”” Thus in the species of Proteo- 
cephalus the position of the vagina with regard to the cirrus-pouch is 
constant, while in the species of Corallobothrium it is inconstant. 

The genus Corallobothrium may be defined as follows: With char- 
acters of family. Scolex bears four suckers situated on anterior surface 
surrounded by many irregular folds and lappets of tissue. Rostellum, 
hooks and spines absent. Neck broad, short. Vagina inconstant in position, 
anterior or poster to cirrus-pouch. Habitat: In Siluridae. Type species: 
Corallobothrium solidum Fritsch. 


CORALLOBOTHRIUM GIGANTEUM NOV. SPEC. 


In life these cestodes are milky-white in color. The scolex is usually 
attached in the region of the duodenum and the proglottids of a mature 
specimen may extend almost to the posterior limit of the intestinal tract 
of the host. They are strongly contractile and may draw themselves up 
to less than half the fully extended length. The strobilization is indis- 
tinct except in the posterior portion of the chain. The body surface is 
excessively wrinkled in this species as is true of the whole genus. The 
proglottids are so firmly joined that considerable force is required to tear 
them apart. The proglottid number varies from 150 to 300 or more. 

Among seventy-five adult individuals before fixation, 60 cm was the 
greatest length observed. After fixation the longest individual was 44 cm. 
The usual length of preserved specimens ranges between 15 and 30 cm. In 
the largest living individuals mature proglottids are about 3 mm long 
by 2.5 mm wide; ripe proglottids 4 to 8 mm long by 1 to 0.5 mm wide, 
according to contraction. In the largest fixed specimen (44 cm) which 
was not fully extended, immature proglottids were from 0.25 to 0.75 mm 
long by 5 mm wide to 1 mm long by 3.5mm wide; mature proglottids from 
1 mm long by 3.5 wide to 2 mm long by 2.25 mm wide; ripe proglottids 


12 ILLINOIS BIOLOGICAL MONOGRAPHS [266 


from 2 mm long by 2.25 mm wide to 5 mm long by 1 mm wide. In an 
alcoholic specimen 22 cm long, containing about 225 proglottids, 3 cm 
from the scolex, the proglottids are about 0.25 mm long by 2.5 to 
2.75 mm wide; about 8 cm from the scolex they are 1 to 1.3 mm long by 
2 to 1.5 mm wide; the last 20 proglottids measure from 1.5 to 2 mm long 
by 1 to 0.5 mm wide. In four toto preparations of sexually mature speci- 
mens which measure from 5 to 15 cm in length, immature proglottids are 
0.10 to 0.84 mm long by 1.51 to 3.10 mm wide; mature proglottids 1 to 
2.2 mm long by 1.3 to 1.89 mm wide; ripe proglottids 1.53 to 3.46 mm 
long by 0.73 to 1.15 mm wide. The segments in the anterior portion are 
always wider than long, but proceeding posteriad the length gradually 
increases at the expense of the width and the posterior proglottids of the 
chain are much longer than wide, the ratio begin from 2:1 to8:1 (Figs 
6, 26). The shape of the cestode in toto recalls that of a whip. 

The form of the scolex is highly variable owing to different states of 
contraction. It may be globose (Fig. 15), quasi-conical (Fig. 5), quadrate 
(Fig. 2), or it may in a measure resemble the head of C. fimbriatum (Figs. 
10, 11). In living specimens it varies from 1 to 2.75 mm in diameter. 
In six alcholic specimens, which are representative of the adult worms, 
the scolex measures from 1 to 2.16 mm wide. The ratio of the width 
to the length varies from about 1:1 to 2:1. Among four toto mounts 
the diameter is 1.47 to 1.68 mm and the length is 0.9 to 1.5 mm. The 
dorso-ventral dimension is usually slightly less than the transverse 
diameter. The four strongly muscular suckers, which are directed an- 
teriorly, are largely concealed beneath a heavy fold of the cortex. From 
this fold project a large number of ridges and a few finger-like processes. 
The folds and lappets are much less pronounced than those of C. fimbria- 
tum and C. solidum (Figs. 5, 10, 15). At times the suckers are drawn in 
and covered entirely (Fig. 11). When this occurs the scolex is drawn back 
into the neck region (Fig. 10). This gives the anterior extremity a flat- 
tened appearance. When the scolex is fully extended, the anterior extremity 
is rounded and the bluntly pointed apex projects from the center (Fig. 
5). There is no rostellum or rudiment of a fifth sucker revealed in five 
sets of frontal and one set of transverse sections. 

The suckers, which measure from 0.45 to 0.59 mm wide and from 
0.53 to 0.59 mm long, vary in shape according to their contraction. In 
some specimens they are nearly spherical while in others they are longer 
than wide. One series of frontal sections shows each pair lying with their 
inner margins touching in the posterior portions while the anterior por- 
tions are quite widely separated, each one making an angle of about 30 
degrees with the longitudinal axis of the worm (Fig. 33). In another 
series of frontal sections, each pair of suckers is separated by a distance 
of about 0.10 mm and they are directed anteriorly so that their longi- 


267] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 13 


tudinal axes are parallel with that of the scolex (Fig. 37). The first series 
was made from a specimen that was fairly well extended, the second 
from one that was much more contracted. The width of the openings 
varies from 0.10 to 0.19 mm and the length of the cavity may be as much 
as 0.33 mm. 

At the apex of the suckers, surrounding the inner half of the opening, 
is a massive set of muscle bands which pass from the margin of the open- 
ing on one side in a circular course to the opposite side of the opening. 
They form a semi-circle about the apex of the suckers. In transections 
they appear as shown in figure 18; in frontal sections they are about 
0.20 mm in diameter and resemble knobs (Fig. 37). Woodland (1925a) 
has reported a similar structure for the suckers of Marsypocephalus rect- 
angulus. This structure functions as a sphincter and, as Woodland has 
suggested, doubtless aids in prehension. It was noted, in dealing with 
the living C. giganteuwm that they were detached with great difficulty 
when the scolex of one fastened itself upon another individual. 

Besides the sphincter, each sucker is encircled by a very compact band 
of muscle fibers 16u thick at the center. These fibers may be divided into 
two groups: those which surround the sucker in the frontal, and those 
which surround it in the sagittal plane. Within this band appears a layer 
of muscle fibers about 0.30 mm thick in the center. These are much less 
compact in their arrangement and all of them extend from the periphery 
of the sucker to the lining of the cavity. Lining the cavity of the sucker 
is a layer of cuticula which measures from 3 to 4u in thickness. 

Anterior to the suckers there is a rhomboid of muscle fibers (Fig. 35) 
and from the margins of the suckers many muscle fibers proceed into the 
neck region (Fig. 37). Posterior to the rhomboid of muscle fibers is a trans- 
verse cross of fibers. At the level of the sphincter a set of fibers pass from 
one sucker to the next (Fig. 18). Transverse sections through the pos- 
terior portions of the suckers show two bundles of muscle fibers, one 
arising from each of the inner margins of the suckers. These bundles 
pass diagonally to a member of the opposite pair. In their course they 
cross each other near the median line to form an eight-rayed cross or 
star (Fig. 28). 

The neck in one set of frontal sections is about 1.60 mm long and 
1.36 mm wide. It is easily recognized in frontal sections but it is not 
evident in much contracted totos. There is frequently very little 
constriction immediately posterior to the scolex. Thus the diameter of 
the neck region, when a specimen is much contracted, may equal that of 
the scolex (Figs. 10, 16, 37). 

The cuticula, which varies from 4 to 9u in thickness, is composed of 
two layers. A thin, more deeply staining layer about 1 thick, covers the 
surface. Beneath it is a thicker, less deeply staining stratum at the inner 


14 ILLINOIS BIOLOGICAL MONOGRAPHS [268 


margin of which is found the very thin basement membrane, less than 
iy thick. Beneath this is a layer of circular muscles about 3u thick. There 
are longitudinal muscle fibers just below the circular muscles. The former 
are interwoven with the outer ends of the spindle-shaped subcuticular 
cells which form a phalanx beneath the subcuticular muscles. These cells 
constitute a layer about 0.03 mm thick. They possess large nuclei which 
vary in shape, some of them being spherical and others elliptical. Each 
cell is drawn out into a fine process which can be followed as far as the 
basement membrane. 

In the strobila the inner longitudinal muscle sheath is exceptionally 
well developed. In mature proglottids it measures from 0.05 to 0.07 mm 
thick. The fibers are arranged in irregularly shaped bundles. Just within 
this layer appears a band of inner circular fibers. This band constitutes 
a stratum from 6 to 16u thick, which encircles the medullary parenchyma. 
In immature segments transverse fibers are quite numerous. Passing 
through the medullary region, as many as 60 bands of dorso-ventral 
fibers were counted in a single section of a mature proglottid. 

The nervous system consists of a ring and two nerve trunks. The former 
is situated in the median region of the scolex at the level of the apices 
of the suckers. In transections it has the form of a cross, the rays of which 
project between the suckers (Fig. 35). Its greatest diameter in adults is 
from 0.33 to 0.40 mm. Arising from the lateral portions of the ring two 
trunks, which measure from 35 to 40u in diameter, pass off parallel to the 
ascending excretory vessel and follow a course between the dorsal and 
ventral pairs of suckers along the inner margin of the longitudinal muscle 
layer (Fig. 30). In the scolex and throughout the proglottids these nerve 
trunks maintain the same relation to the inner longitudinal muscle sheath, 
and their diameter varies but little at any point in the chain. 

The excretory tubules are very prominent in sections of this species. 
Four principal tubules are easily seen in sections of proglottids which 
are immature. When the reproductive organs are fully developed the 
excretory system is so crowded that at times it is difficult to distinguish 
it clearly. The four main longitudinal trunks are located in the lateral 
regions of the medullary parenchyma roughly parallel to the longitudinal 
nerve trunk of each side, the descending lying nearer the center than 
the ascending vessels. 

The ascending tubules, which measure from 12 to 18u in diameter, 
lie only 5 to 15u from the nerve trunk, both in the neck region and in the 
immature proglottids. In these regions they follow a straight course with 
only slight deviations. The descending tubules are much larger in the 
regions just named, as they measure from 24 to 40u. Their course is much 
more tortuous and their position with reference to the nerve trunk is 
much less definite. 


269] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 15 


The structure of the ascending and descending trunks is very different. 
Transections of the former showa heavy wall of hyaline material about 
1p thick surrounded by what appears to be a single layer of muscle fibers. 
Also the parenchymal cells are compactly arranged about these tubules 
(Fig. 40). The descending trunks lack the hyaline wall and also the layer 
of muscle fibers, and they are surrounded by a less compact arrangement 
of the parenchymal cells. 

In a larval specimen about 4 mm long, cut in transverse section, the 
ascending trunks arise 0.14 mm from the posterior end of the individual 
where they measure from 2 to 5uin diameter. They are found in the lateral 
medullary parenchyma. As they proceed anteriorly they increase slightly, 
measuring 8 to 10u at a point about 0.8 mm from the scolex. As they 
enter the scolex they measure about 8u in diameter. Upon reaching the 
scolex they pass laterally between the suckers and the longitudinal muscle 
layer of the scolex, following the outer margins of the suckers until the 
anterior level of the latter is reached, when they proceed inward toward 
the median line where they follow a tortuous course between the suckers 
before uniting with the descending trunks. 

Branches of the descending trunks are more numerous in the central 
region of the scolex. The apex is ramified with both ascending and descend- 
ing branches, but about the bases of the suckers there occur numerous 
coils of the descending tubules. Upon reaching the neck region, the main 
vessels proceed laterally, the ascending running dorsal and the descending 
ventral to the nerve trunk. In the neck the ascending trunks measure 
about 0.02 mm, or twice the diameter of the ascending ones. Through- 
out the length of the individual there is little difference in the diameter 
of the descending tubules since they measure from 20 to 25y in the pos- 
terior region. There are no cross trunks apparent between the vessels 
of the same or opposite sides in the parasites at this stage of develop- 
ment. Furthermore, they follow practically a straight course throughout. 
About 0.08 mm from the posterior end, the two descending trunks empty 
into an excretory bladder. Just below the point of union with the de- 
scending tubules, the bladder measures 20u dorso-ventrally and 27 trans- 
versely. It narrows posteriorly and empties through a small duct at the 
extreme posterior end of the individual. 

Frontal sections were made of a strongly contracted individual, about 
15 mm long, in which a large number of proglottids had been differentiated. 
Here the excretory system is much more highly developed than in the 
individual just discussed. The position of the ascending and descending 
trunks with reference to each other has been slightly altered. At this 
stage they are parallel in a horizontal plane, with the ascending trunk 
more lateral or nearer the longitudinal nerve on each side. The diameter 
of the ascending is less than that of the descending trunks throughout 


16 ILLINOIS BIOLOGICAL MONOGRAPHS [270 


their extent. Both are more or less coiled in this individual. With the 
differentiation of proglottids an addition to the descending system has 
occurred. At the posterior limit of each proglottid is found a cross-trunk 
which in many instances equals in diameter the descending tubules. 
Also a small duct, 8u in diameter, arises near the confluence of the des- 
cending and cross trunks. This passes to the posterio-lateral margin of 
the proglottids and empties to the outside through a small duct and pore 
(Figs. 34, 38). These same features are found also in the fully mature 
segments, but owing to the pressure of the reproductive organs they are 
more difficult to distinguish. The excretory system of this species agrees’ 
in all essential features with the description of Riggenbach (1896) for 
Ephedrocephalus lobosus (=Corallobothrium lobosum). 

A study of frontal sections of the scolex and strobila in an extended 
and contracted condition has lead me to the conclusion that this system 
has more than an excretory function. When the scolex and strobila are 
contracted, the vessels are greatly diminished (Figs. 16 and 38). When 
the opposite is the case, the vessels are much distended (Figs. 21 and 34). 
This suggests the probability that the excretory vessels aid in the exten- 
sion of the scolex and strobila. 


Reproductive Organs 


A common genital sinus, which measures about 64 in diameter? 
occurs on the lateral margin of the proglottid from one-quarter to one~ 
half the proglottid length behind the anterior margin. The usual position 
is within the anterior one-quarter of the proglottid. It is irregularly 
alternate in successive segments. The cortical wall surrounding the sinus 
protrudes sufficiently to be seen with the naked eye in toto mounts. 
This constitutes a genital papilla. The character and arrangement 
of the reproductive organs, for the most part, is identical with that of 
the other Proteocephalids. Each sexual unit occurs within the inner 
longitudinal muscle sheath (Fig. 20). 

The male system will be considered first. The testes vary considera- 
bly in shape. Some of them are almost spherical, others pear-shaped, 
but the majority are elliptical. They measure from 0.05 mm to 0.08 mm 
in length, and from 0.03 to 0.06 mm in diameter. The number present 
in a single proglottid is from 80 to 100. They lie in a continuous field 
between the vitelline glands, extending from the anterior margin of the 
proglottid posteriad to the level of the ovary. Between the vitelline 
glands and the uterus they occur in two or three layers, but dorsal to the 
uterus only a single layer is present. Vasa efferentia were not observed 
in any of the preparations. In many sections the vas deferens passed 
among the testes and often there appeared to be a connection between 
the walls of the two. This relation could not be established for a sufficient 
number to warrant the statement that such is the usual condition. 


271] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 17 


At the level of the cirrus the thin-walled vas deferens, which measures 
from 0.03 to 0.05 mm in diameter, forms a compact mass of 15 to 20 
coils that almost completely fill the medullary space between the vitel- 
laria and uterus and extend anteriad and posteriad from 0.25 to 0.33 mm. 
(Figs. 7, 26). In mature proglottids they are usually distended with 
spermatozoa. At the point of entrance into the cirrus-pouch the vas 
deferens ends abruptly and at this point the ductus ejaculatorius takes 
its origin. After describing two or three coils it passes into the cirrus. 
The duct at the point of entrance into the cirrus-pouch measures about 
16u. A short distance beyond its point of entrance it doubles in diameter. 
Upon entering the cirrus it narrows again. Its lumen varies from 8 to 12u 
in diameter. Its structure is almost identical with that of the cirrus. 

The cirrus-pouch is usually elongate-oval in shape when the cirrus is 
inverted, but its form varies when the cirrus is everted. Apparently the 
extent of protrusion of the cirrus affects the shape of the cirrus-pouch. 
It assumes a variety of forms in successive segments. Some appear wedge- 
shaped, while others recall the form of a gourd (Fig. 7, 26). When the 
cirrus is inverted, the pouch measures from 0.26 to 0.33 mm long by 0.09 
to 0.13 mm in greatest diameter. The wall of the pouch is from 3 to 5y 
in thickness and is composed of longitudinal muscle fibers. Between the 
wall of the pouch and the cirrus is a loose network of connective tissue fibers 
with scattered nuclei. Immediately surrounding the cirrus is a compact 
layer of cells about 8u thick. Wagner (1917) in his description of Proteo- 
cephalus torulosus calls a group of cells similarly located, subcuticular 
cells. Benedict (1900) in his description of P. filicollis and P. amblop- 
litis designates them as gland cells. 

When the cirrus is inverted it measures from 0.20 to 0.23 mm in length 
and about 0.02 mm in greatest diameter and about the same at its point 
of union with the ductus. It is roughly cone-shaped, the base of the cone 
being adjacent to the genital sinus. In transections the wall of the cirrus 
shows a series of closely applied folds which almost close the lumen. The 
inverted cirrus is lined with a cuticular layer that extends to the ductus. 
Surrounding this layer are found longitudinal and circular muscles. 
When completely everted the cirrus reaches a length of 0.5 mm. Its dia- 
meter just outside the genital sinus is about 0.1mm, while its distal por- 
tion has a diameter of about 0.05 mm (Fig. 17). 

In the female system the vagina opens into the genital sinus beside 
the cirrus in the same horizontal plane. Frontal sections of two indi- 
viduals showed that the vagina may be either anterior or posterior to the 
cirrus. In one individual the vagina opened anterior to the cirrus on the 
right side in 14 proglottids, and on the left side anterior to the cirrus, in 
12 proglottids. It opened posterior to the cirrus on the right side in 10 
proglottids and posterior to the cirrus on the left side also in 10 proglottids. 


18 ILLINOIS BIOLOGICAL MONOGRAPHS [272 


In another specimen the relation was as follows: anterior to cirrus, right 
side, 18; anterior to cirrus, left side, 20; posterior to cirrus, right side, 10; 
posterior to cirrus, left side, 13. In these two specimens there is a close 
approach to equality in the dextral and sinistral position, and antero- 
posterior relation of the vagina and cirrus. For a distance of about 0.10 
mm from the sinus the diameter of the vagina may almost equal that 
of the cirrus, and like the latter has a lining of rather heavy cuticula. A 
sphincter vaginae is present but weakly developed. At the end of the 
distance just mentioned the vagina narrows, and from there to the semi- 
nal receptacle it measures from 11 to 24y and its lumen varies from 6 to 
204 in diameter. The greater diameter is only reached when the vagina 
is distended with spermatozoa. Since the vaginal opening may lie either 
anterior or posterior to the opening of the cirrus, the course of the vagina 
will vary somewhat. From the level of the genital sinus it may pass an- 
teriorly for a short distance before proceeding toward the median line, 
or it may run between the coils of the vas deferens in a direct course to- 
ward the median line of the segment. At the same time it is directed 
ventrad. It passes through the ventral region of the medullary cortex 
for a short distance, then turns dorsally and for the remainder of its course 
to the seminal receptacle, it lies above the uterus. It is found in this 
last position throughout about two-thirds of its length. Anterior to 
the ovarian commissure when the proglottid is much extended, and 
posterior to it in contracted segments, the vagina widens gradually for about 
0.10 mm, then it narrows suddenly, giving rise to a pear-shaped seminal 
receptacle which equals the width of the vagina at ifs narrowest point 
and is about 0.04 mm at its widest portion (Fig. 23). Leading from this 
structure is a narrow duct, the lower vagina, which joins the oviduct. 
This portion may be straight (Fig. 23) or coiled (Fig. 39). The position 
of the duct is dependent upon the contraction of the proglottids. Aside 
from a cuticular lining for a short distance from the genital sinus, the 
wall of the vagina is composed of muscle fibers and measures from 3 to 
5u in thickness. Only longitudinal fibers were distinguished. Like the 
cirrus, it is surrounded by a layer of gland cells throughout its length. 

The bi-lobed ovary lies in the posterior portion of the proglottids. 
In fully extended ripe proglottids it is H-shaped. The two wings are broad 
and bluntly rounded posterior to the commissure, but anterior to it they 
taper to a rather sharp point and measure from 0.86 to 1.32 mm in length. 
The width of the ovary at the level of the commissure is from 0.33 to 
0.46 mm (Figs. 23, 26). In fully extended ripe proglottids the structure 
of the ovary has a latticed or corded appearance as it is made up of a 
network of thin-walled tubules which are interwoven with one another. 
In mature proglottids the lobes of the ovary are much shorter and more 
compact, measuring from 0.46 to 0.80 mm in length and from 0.75 to 
1.06 mm in width at the level of the commissure (Fig. 26). 


273] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 19 


As revealed by sections, the two lobes lie just beneath the inner longi- 
tudinal muscles in the dorsal region of the medullary parenchyma but the 
commissure dips ventrally. Near the median line, it connects with the 
muscular oocapt which is spherical in shape and measures about 20y in 
diameter. It is surrounded by a layer of gland cells about 4u thick. From 
the oocapt in mature proglottids the oviduct usually changes direction 
three or four times before reaching the posterior margin of the segment 
where it turns laterad and dorsad. Near its lateral limit it empties into 
the ootype. The greatest diameter of the oviduct is about 24u. The lower 
vagina joins the oviduct at its last bend before the ootype is reached 
(Fig. 39). 

The ootype, which is about the same diameter as the oviduct, passes 
laterally and dorsally for a short distance and then proceeds anteriorly. 
Near the anterior level of the vitelline receptacle it empties into the uter- 
ine passage. The wall of the ootype is very similar to that of the oviduct. 
The only striking difference between the two structures is the presence 
of the shell gland about the former, some cells of which measure 48 long 
by 16u wide. The processes of these cells pass into the wall of the ootype. 

The uterine passage, which leads from the ootype, measures about 16 
in diameter. After bending upon itself once or twice it passes dorsal 
to the vagina and empties into the uterus near the median line ata point 
near the middle of the proglottid. 

In mature proglottids the uterus has a diameter equal to about half 
the proglottid width. It is separated from the vitellaria on each side 
and from the dorsal musculature by the testes. It extends from the an- 
terior margin to near the level of the ovarian commissure. It opens to 
the exterior through two or three uterine pores near the ventral median 
line. In mature proglottids the uterus possesses from 10 to 15 lateral 
pouches (Fig. 26). Their beginnings are very distinct in immature pzo- 
glottids. In fully extended, partially spent proglottids the lateral pouches 
are relatively greatly reduced, giving the uterus the character of a tube 
with irregular borders (Fig. 6). 

The vitellaria form two columns of a diameter of 0.08 mm which 
extend from the anterior to the posterior margin of the proglottid. They 
crowd the longitudinal nerve trunks close to the inner muscle sheath, 
They are follicular in structure, the follicles emptying into a central 
tubule. Near the level of the ovarian commissure a duct 16u in diameter 
leads inward from each side. The two unite anterior to the oocapt, form- 
ing a common vitelline duct which proceeds posteriorly for about 0.05 mm 
and then widens to form the vitelline reservoir, which is from 40 to 56u 
Jong and from 28 to 36u wide. From the reservoir a narrow duct leads 
into the ootype near its point of union with the oviduct (Fig. 39). 


20 ILLINOIS BIOLOGICAL MONOGRAPHS [274 


AMPHITYPY 


Amphitypy, or complete organ reversal, occurs in the position of all 
the interovarian organs in successive segments. This condition is not 
correlated with the position of any of the other organs of the reproductive 
system. For example, the genital sinus may be on the left margin with 
the vagina anterior to the cirrus and with the interovarian organs dis- 
posed as shown in figure 39. In another segment the genital sinus will 
be on the right with the vagina posterior to the cirrus and the 
interovarian organs will be arranged the same as in the figure just cited, 
or they may be just the reverse. The following data were taken from 
seven successive segments cut in frontal section. 


Right | Right} Left | Right} Right) Left | Right 


Genital sinus 


Vagina anterior to cirrus + AF 

Vagina posterior to cirrus” + + + + oh 
Oviduct right + + + oF 

Oviduct left + 
Seminal receptacle right + oF 


Seminal receptacle left 
Vitelline reservoir right 
Vitelline reservoir left 


Amphitypy has been observed among the Trematoda by many in- 
vestigators (Looss, 1902: 789) but I have not discovered any reference to 
such a condition among the Cestoda. 

A description of the living eggs is given later in this paper conse- 
quently only the ova within the uteri of sectioned specimens are considered - 
here. The outer egg covering is so variable in amount and so irregular 
in shape that it is difficult to distinguish in preserved material. The 
second membrane is spherical and measures from 14 to 19u in diameter. 
This membrane is closely applied to the oncosphere which measures 
from 8 to 13u in diameter. The eggs of this species are the smallest pro- 
duced by any of the species of Corallobothrium thus far described. 

Because of the tremendous difference in the length as compared with 
the other two species, the smaller number of testes, and the great dif- 
ference in the egg measurements, besides the other distinct characters 
to be pointed out, I regard the parasite under consideration as a new 
species which I shall designate as Corallobothrium giganteum. This name 
was given not only because it is the largest species of its genus but also 
because it is one of the largest known cestodes infesting fresh-water fish. 


Max. Sucker No. of Second egg Size of 

length sphincter testes membrane oncosphere 
C. solidum 4 cm. absent 140 to 180 20 to 24u 13 to 16u 
C. giganteum 44 cm. present 80 to 100 14 to 19u 8 to 13u 


C. fimbriatum 8 cm. absent 100 to 125 28 to 36u 16 to 24y 


275] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 21 


Only a superficial examination is necessary to separate this species 
from Corallobothrium solidum or C. fimbriatum. Besides the points con- 
sidered in the preceding comparison, C. gigantewm is distinct from the other 
species in the character of the scolex (Figs. 5, 10, and 15), the size and shape 
of the proglottids, shape of the ovary, and in the less extensive develop- 
ment of the uterus. The measurements in the preceding comparison 
were made on preserved material. 


CORALLOBOTHRIUM FIMBRIATUM NOV. SPEC. 


This cestode is usually found in the anterior portion of the intestine 
of Ictalurus punctatus, Leptops olivaris, or Ameiurus melas. The length, 
when ripe proglottids are present, varies from 15 to 80 mm among pre- 
served specimens. The maximum breadth reaches 2.6 mm. Living in- 
dividuals frequently reach a length of 70 to 90 mm when well extended. 
The strobilization is very distinct and the segments are more easily de- 
tached than those of C. giganteum. The number of proglottids ranges 
from 40 to 90. They vary in shape and size according to the state of con- 
traction and the body size of the individual. The following measurements 
in millimeters were taken from 6 prepared specimens: 


Immature proglottids 
length 0.09, 0.16, 0.41, 0.19, 0.53, 0.16, 0.31, 0.27, 0.21 
breadth 0.78, 0.46, 2.40, 1.57, 1.89, 1.68, 1.36, 1.15, 0.56 
Mature proglottids 
length 1.00, 0.95, 1.05, 0.50, 0.63, 0.73, 0.73, 0.63 
breadth 1.9, 1.26, 1.36, 0.59, 1.26, 1.26, 2.60, 1.05 
Ripe proglottids 
length 1.05, 1.05, 1.01, 1.05, 1.00, 0.94, 1.05, 1.36, 1.36, 0.84 
breadth 1.78, 2.60, 1.01, 1.01, 1.20, 1.36, 1.05, 0.73, 0.73, 0.84 


The thickness varies from 0.5 to 0.7 mm. The immature and mature 
proglottids are almost invariably broader than long, while the ripe seg- 
ments are longer than broad. In transections some ripe proglottids are 
flattened dorsally but rounded ventrally; in others the reverse is true. 

In the descriptions of C. solidum, with which this form agrees very 
closely, that of Fuhrmann (1916) is the only one containing adequate 
measurements. He records the length of first proglottids as 0.09 mm; 
5 mm behind the scolex as 0.23 mm and posterior segments as 0.65 mm. 
He gives the same maximum length and breadth as did Fritsch (1886) 
since he used the latter’s preparations. Janicki (1926) gives the maximum 
length as 60 mm and maximum breadth as 3.5mm but fails to record 
any other measurements for the proglottids. I have been unable to find 
in any C. jfimbriatum specimens the definite surface structure which 
Janicki describes and figures for C. solidum. He states, “Von den zwei 


22 ILLINOIS BIOLOGICAL MONOGRAPHS [276 


mir vorliegenden Exemplaren weist das grdéssere eine sehr eigentiimliche 
Gestaltung der Kérperobeflache auf, indem die Rindenschicht durch tiefe 
longitudinal wie transversal verlaufende Einkerbungen das Bild einer mosai- 
kartigen Tafelung hervorruft.’”’ There are, to be sure, longitudinal and 
transverse grooves which are largely the result of contraction (Fig. 1), 
but they are never as regular in occurrence as Janicki indicates for C. 
solidum. 

This species possesses the same type of scolex as C. solidum and Goe- 
zeella silurt Fuhrmann 1916. It measures from 1.26 to 3.75 mm in trans- 
verse diameter The latter measurement was found in only one case and 
then the scolex was completely expanded. Figure 4 represents the scolex 
in a more expanded condition. Among 15 individuals bearing ripe pro- 
glottids the scolex usually measured about 2 mm in transverse diameter. 
The dorso-ventral is always less than the transverse diameter. In one 
fully expanded scolex the former is 2.31 mm and the latter is 2.61 mm. 
It is evident that the scolex of this species is subject to such ex- 
tremes that many individuals must be studied to establish the 
range in size. When fully expanded, the scolex appears as a disc surmount- 
ing the proglottids and is set off sharply from the neck. In some con- 
tracted specimens the scolex is not thus sharply set off (Fig 1). The 
collar-like structure which surrounds the suckers may be folded over 
them laterally and dorso-ventrally, thus concealing them from view. 
Extending from each of the suckers toward the periphery of the scolex 
are deep grooves which mark the limits of the folds just described. The 
anterior surface of the scolex thus frequently resembles that of Ephed- 
rocephalus lobosus as figured by Riggenbach (1896, Pl. 8, fig. 23b). The 
apex may be protruded as shown in figure 22, or it may be invaginated 
as indicated by figure 19. A rostellum is lacking, and hooks of any kind 
are absent. 

Four suckers are situated in the anterior surface of the scolex. Their 
form and size vary according to the state of contraction. The following 
measurements in millimeters were made on sectional material: 

Length 0.36, 0.43, 0.41, 0.37, 0.37, 0.38, 0.33, 0.45, 0.33, 0.53 
Diameter 0.56, 0.53, 0.51, 0.37, 0.41, 0.40, 0.41, 0.38, 0.43, 0.60 
Diameter of opening 0.13, 0.19, 0.21, 0.06, 0.06, 0.06, 0.12, 0.18, 0.18, 0.23 


The measurements which follow were taken from toto-mounts: 
Length 0.30, 0.23, 0.23, 0.19, 0.26, 0.39, 0.51 

Diameter 0.38, 0.26, 0.26, 0.23, 0.23, 0.53, 0.88 

Diameter of opening 0.16, 0.09, 0.06, 0.11, 0.13, 0.13, 0.41 


A description of the musculature of the scolex and its suckers is un- 
necessary, since there is no apparent difference in this regard between 
C. fimbriatum and E. lobosus as described by Riggenbach. The peculiar, 


277] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 23 


knob-like structure or sphincter found in the suckers of C. giganteum is 
wanting here. In the three descriptions of C. solidum the size of the 
suckers is not given. From Fritsch’s figure which was drawn to scale, I 
estimated the length of the suckers as 0.6 mm and the width as the same. 

A neck is present, but its dimensions are dependent upon the state 
of contraction and the size of the specimen. Among eight individuals 
the neck in three was longer than wide, but in five cases it was wider 
than long. In millimeters the necks measure as follows: 

Length 0.46, 0.63, 0.46, 0.53, 0.72, 0.96, 1.05, 1.26 

Breadth 0.33, 1.89, 0.36, 0.56, 0.86, 1.21, 1.57, 0.94 


In his description of C. solidum Fritsch states, ‘‘Man erkennt ohne 
Schwierigkeit, dass die schmalen, ohne Vermittelung eines Halses dem 
Kopf angefiigten Glieder, sehr bald geschlechtsreif werden.”’ Fuhrmann, 
using the same preparations, reports a short neck, while Janicki states 
that no neck is present in the specimens examined by him. 

The cuticula of this species varies from 5 to 9u in thickness which 
corresponds very closely to that of Corallobothrium giganteum. It is 
divided into two layers, an inner which stains deeply with hematoxylin, 
and an outer layer which stains very little. Beneath the cuticula courses 
the very thin basement membrane (less than 1u). The subcuticular 
musculature shows a variation from the usual condition since the longi- 
tudinal fibers, which are present in a single layer, lie closely applied to 
the basement membrane. The usual layer of circular fibers was not dis- 
tinguishable. Beneath the muscle fibers just described occur the spindle- 
shaped subcuticular cells which constitute a layer about 0.03 mm thick. 
The cortical parenchyma is very loosely constructed. In some sections 
it reveals a network of rounded spaces which measure from 6 to 35y. 

The musculature of this form is highly developed, but less so than 
in C. giganteum. Separating the cortical and medullary parenchyma are 
found the inner longitudinal muscles which constitute a layer from 16 to 
30u thick. These fibers are arranged in irregularly shaped bundles, but 
the bundles are not grouped in layers (Fig. 27). In C. solidum both Fuhr- 
mann and Janicki report that laterally the muscles are more highly 
developed. This is also the case in C. fimbriatum. In this species trans- 
verse bundles, within the longitudinal muscle layer, are very numerous 
in immature proglottids; but in mature segments they are crowded against 
the longitudinal bundles by the growth of the reproductive organs. Many 
dorso-ventral bundles are present between the pouches of the uterus and 
in the lateral margins of the medullary parenchyma. 

The nervous system consists of a nerve ring situated in the apex of 
the scolex near the anterior level of the suckers. A nerve trunk which 
measures 0.03 mm in diameter arises from each side of the ganglion and 


24 ILLINOIS BIOLOGICAL MONOGRAPHS [278 


passes posteriad between the outer margins of the two pairs of suckers. 
These trunks extend throughout the strobila in the lateral margin of the 
medullary parenchyma close to the longitudinal muscle sheath, as in 
C. giganteum and most Proteocephalids. 

The excretory system is highly developed, but less so than in C.gigan- 
teum. It represents the typical condition as there are two pairs of vessels, 
a dorsal or ascending pair, and a ventral or descending pair. The former 
are more lateral in position and are found throughout the strobila closely 
applied to the nerve trunks. Their course is much less tortuous than 
that of the descending vessels. No branches could be found issuing from 
them. In ripe proglottids they measure about 4y in diameter, but pro- 
ceeding anteriorly the diameter increases until in the neck region it may 
attain as much as 24u. The structure of the wall of the ascending trunk 
is very similar to that described for C. giganteum (Fig. 40). Frequently 
in ripe proglottids the vitellaria surround the dorsal vessel and it might 
be mistaken for a longitudinal vitelline duct. 

The descending trunks, as already mentioned, describe a tortuous 
course. Their position is less rigidly fixed than that of the ascending 
vessels, because of the crowding of the reproductive organs, but they 
are always found in the ventral medullary parenchyma proximal to the 
ascending vessels. Their diameter varies from 24 to 40u. Their walls 
are much thinner than those of the ascending vessels. The cross com- 
missure which connects the descending vessels and the small vessels 
leading to the exterior which have been described for C. giganteum, are 
wanting in C. fimbriatum. Likewise I have been unable to discover any 
such vessels as Janicki has described for C. solidum. He states: ‘Die 
Foramina secundaria gelangen im hinteren Teil der Proglottis auf der. 
ventralen Seite zur Entwicklung und stehen vermittelst besonderer 
Zweiggefissen mit den zwischen den grossen Ventralstimmen sich 
ausspannenden und vielfach secundir anfgelésten Commissuren in Ver- 
bindung.” At no point can I find any vessels passing beyond the inner 
ongitudinal muscle sheath. The behavior of the ascending and descend- 
ing vessels in the scolex is almost identical with that already described 
for C. giganteum. 


Reproductive Organs 


The common gential sinus occurs irregularly alternate on the lateral 
margin of the proglottid, almost invariably within the anterior fourth 
of the segment. A genital papilla is lacking in this species. The character 
and arrangement of the reproductive organs is typically Proteocephalid, 
as all the sex organs are contained within the inner longitudinal 
muscle sheath. 


279] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 25 


In immature proglottids where the testes are not crowded upon them- 
selves or upon other organs, they are typically spherical and measure 
from 0.02 to 0.04 mm in diameter. In the mature proglottids, their shape 
varies from spherical to elongate oval, the form being dependent on the 
degree of pressure to which they are subjected (Fig. 32). The spherical 
testes measure from 60 to 72y in diameter while the others range from 
64 long by 48 wide to 80u long by 724 wide. The number of testes 
present in mature proglottids ranges from 100 to 125. Before the uterus 
has undergone a great deal of development the testes occupy all the 
available space in the medullary parenchyma (Fig. 14), but as the uterus 
grows it crowds the testes dorsally and laterally (Fig. 31). In mature 
proglottids only a single layer is present dorsal to the uterus, and only 
two or three layers are usually found lateral to it. In ripe proglottids the 
testes are crowded out of the lateral portions of the medullary parenchyma 
by the uterus, which extends to the vitellaria on each side (Fig. 31). 
They are found between the uterine pouches and frequently in the ven- 
tral portion of the medullary parenchyma. In the fully ripe segments 
they have almost completely disappeared. 

Vasa efferentia were not distinguishable in any of the preparations. 
The vas deferens which measures from 16 to 50u in diameter is extremely 
well developed. Its coils, from 25 to 30 in number, extend from the mid-line 
of the segment to the cirrus-pouch (Fig. 14). In immature segments 
they occupy nearly the anterior fourth of the medullary parenchyma, 
but when the segments become mature they are forced dorsally or between 
the pouches of the uterus (Fig. 29). Spermatozoa are present in the 
vas deferens when the uterus is in a very early stage of development 
(Fig. 14), ie., in proglottids which are otherwise immature. Upon reach- 
ing the cirrus-pouch the vas deferens passes over into the ductus ejacu- 
latorius. In immature segments it proceeds to the cirrus without coiling 
but in mature and ripe proglottids it describes three or four coils before 
emptying into the cirrus. Its diameter, upon entering the cirrus-pouch 
is about 16u, but it widens as it proceeds, and frequently it may reach a 
diameter of 30 to 40u before emptying into the cirrus. As it enters the 
cirrus it narrows again to about 16y in diameter. 

When inverted, the cirrus measures from 0.13 mm long by 25u wide 
to 0.19 mm long by 36 wide. It is typically club-shaped, with the narrow 
end at the proximal portion of the cirrus-pouch. When everted the cirrus 
measures from 0.16 to 0.23 mm in length and from 40 to 44 in greatest 
diameter. It tapers gradually from its proximal to its distal end, where 
it measures about 0.02 mm in diameter. .A terminal dilatation is wanting 
(Fig. 36). The cirrus-pouch varies from elongate oval, when the cirrus 
is inverted, to gourd-shape, when the cirrus is everted. It measures 
from 0.17 to 0.24 mm in length and from 0.07 to 0.09 mm in greatest 


26 ILLINOIS BIOLOGICAL MONOGRAPHS [280 


diameter. None of the descriptions of C. solidum record the size of the 
cirrus inverted or everted. The cirrus-pouch according to Fuhrmann is 
pear-shaped and measures 0.5 mm long by 0.22 mm wide. Janicki makes 
no reference to the shape but reports the length as 0.34 mm. The struc- 
tural details of the vas deferens, cirrus and cirrus-pouch do not vary 
from those described for C. giganteum. 

As in C. giganteum, the vagina opens into the genital sinus beside 
the cirrus in a horizontal plane, and may be either anterior or posterior 
to the latter. Both the cirrus and vagina pass into the medullary paren- 
chyma between the dorsal and ventral excretory vessels and ventral to 
the longitudinal nerve trunk. 

From the genital sinus the vagina passes along the ventral margin 
of the medullary parenchyma for about one-fourth of the proglottids 
diameter; then it courses dorsally and posteriorly to the mid-line of the 
proglottid, which it follows to the level of the ovarian commissure (Fig. 
29). The course of the lower vagina will be discussed presently. 

A sphincter vaginae is absent, or only weakly developed. The first 
part of the vagina, or the portion that usually extends from the genital sinus 
to about the mid-line of the segment, measures from 11 to 16u in diameter. 
Its walls are somewhat heavier than those of the succeeding portion. The 
second portion of the vagina is thin-walled, and frequently reaches 32u 
in diameter, but in such cases it is distended with spermatozoa. This 
portion of the vagina functions as a receptacle for spermatozoa. A dis- 
tinct seminal receptacle, such as is found inC. giganteum is not present. 
Fuhrmann states that a seminal receptacle is wanting in C. solidum. Near 
the level of the ovarian commissure the second portion of the vagina is 
greatly reduced in diameter, giving rise to the narrow, more muscular, 
lower vagina which measures from 8 to 124 in diameter. It passes dorsal 
to the ovarian commissure and follows along the lateral margin of the shell 
gland. Near the posterior margin of the segment it empties into the 
oviduct (Fig. 41). 

The bi-lobed ovary lies in the posterior portion of the proglottids 
midway between the dorsal and ventral longitudinal muscles. In some 
transections the ovary presses against the dorsal and ventral longitudinal 
muscles thus filling much of the medullary space in the posterior end of 
the segment. Its shape may be roughly omegoid (Fig. 9) or it may be 
pyramidal (Fig. 8). The contraction of the segment affects the shape 
very materially. When much contracted, each wing of the ovary shows 
several secondary lobes (Fig. 12), which are less evident when the seg- 
ment is fully extended (Fig. 13). A commissure joins the inner anterior 
margins of the two wings of the ovary. The inner posterior margins pass 
around the shell gland and proceed toward the median line but do not 
join. From the shell gland the two wings extend laterally. The pos- 


281] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 27 


terior margin of each wing parallels the septum of the segment, but the 
anterior margin dips down gradually until near the vitellaria on each 
side the two margins meet in a rather acute angle. Figures 8, 9, 12 and 13 
represent some of the variations in the shape of the ovary. In an ex- 
panded ripe segment 1.58 mm wide the ovary measures 0.79 mm in dia- 
meter and only 66y at the point of greatest length. In an expanded mature 
segment 1 mm wide the ovary measures 0.56 mm wide and 0.01 mm 
through the region of greatest length. 

The following measurements in millimeters were made on mature 
segments which were more or less contracted: 

Width of segment: 1.15, 1.11, 1.19, 1.26, 1.26 

Width of ovary: 0.45, 0.46, 0.53, 0.54, 0.53 

Length of ovary: 0.22, 0.23, 0.24, 0.21, 0.23 
These measurements indicate that the width of the ovary is a little less 
than half the proglottid diameter. 

Near the median line, and on the ventral surface, the commissure 
empties into the oocapt which measures from 28 to 38y in diameter and 
is surrounded by a layer of gland cells about 16 thick (Fig. 29). Fuhr- 
mann gives the diameter of the oocapt of C. solidum as 28u. From the 
oocapt the oviduct, whose maximum diameter is about 24u, proceeds 
along the lateral margin of the interovarian space. Just before reaching 
the intersegmental septum it changes direction and crosses the inter- 
ovarian space. Near the opposite side it bends posteriad and dorsad. 
After receiving the lower vagina it passes along the posterior margin of 
the proglottid. Upon reaching the middle of the interovarian space it 
turns anteriorly. Near this point it receives the vitelline duct and empties 
immediately into the ootype (Fig. 41). 

The ootype, which measures from 12 to 16u in diameter, lies in the 
dorsal portion of the medullary parenchyma. It is surrounded by the 
shell gland which is highly developed and measures from 0.09 to 0.16 mm. 
in diameter (Fig. 32). From its connection with the oviduct the ootype 
continues anteriorly from 0.09 to 0.10 mm, then it empties into the ute- 
rine passage. The uterine passage is a very thin-walled tube measuring 
from 16 to 20u in diameter. From the ootype it proceeds anteriorly 
and dorsally. After making several coils it empties into the uterus near 
the middle of the proglottid. 

The uterus begins growth in the ventral region of the medullary 
parenchyma but as it develops takes up more and more space, until 
in the ripe proglottids it crowds the vitellaria, excretory vessels and longi- 
tudinal nerves at each side, and occupies all possible space (Figs. 25, 31). 
The testes, vas deferens and other organs are crowded between the ute- 
rine pouches or flattened against the longitudinal muscle layer. The 
ovary is frequently pressed against the posterior limit of the segment 


28 ILLINOIS BIOLOGICAL MONOGRAPHS [282 


to such an extent that it is distorted in shape and may be difficult to 
distinguish (Fig. 25). At this stage the proglottid is hardly more than 
an egg-sac. From the main uterine stem arise from 10 to 14 uterine pouches, 
each of which in turn produces from 2 to 5 secondary pouches (Figs. 
29, 32). On the ventral surface of the ripe proglottids occur one or two 
uterine pores. 

The vitellaria form two lateral columns which extend from the an- 
terior to the posterior region of the segment. They are more dorsal than 
ventral in position. The ventral excretory vessels lie beneath them. Their 
transverse diameter, whichis dependent on theamountof contraction, ranges 
from 0.06 to 0.19 mm. At the level of the ovary or near the posterior 
limit of the proglottid each column turns inward until it comes in con- 
tact with the wings of the ovary (Fig. 14). From these inward-directed 
portions arise two vitelline ducts, one from each side, which run ventrad 
and meet lateral to the oocapt, forming the common vitelline duct, this 
proceeds dorsally and empties into the oviduct near its union with the 
ootype. The diameter of the two ducts just mentioned depends on the 
presence of vitelline cells; when empty they are indistinguishable, but 
when they contain vitelline cells the lumen measures from 11 
to 16u. At the point where the lateral ducts empty into the common 
vitelline duct a diameter of 32y is frequently attained. From this region 
to its union with the oviduct the common vitelline duct measures only 
from 16 to 24y in diameter. A distinct vitelline reservoir, like that in 
C. giganteum, is lacking in this species (Fig. 41). The vitelline cells pre- 
sent in the ducts are ovoid in form and measure from 11 to 14y through 
their long axes. 

This species shows amphitypy, the same irregularity in the arrange- 
ment of the organs of the interovarian space as described for C. gigan- 
teum; that is, in successive segments a complete reversal occuts. This 
reversal is not correlated with the right or left position of the genital 
pore, nor with the position of the vagina with reference to the cirrus. 
The organs may be arranged as shown in figure 41 or they may be just 
the reverse. In five successive segments the arrangement of the organs 
as shown below was observed. 


Left Left 


of 


Genital Pore 


Vagina anterior 
Vagina posterior 
Oviduct right 
Oviduct left 
Vitelline duct right 
Vitelline duct left 
Lower vagina right 
Lower vagina left 


+ 
+ 


283] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 29 


A description of the living egg of the species is given later. Only the 
interuterine eggs of prepared specimens will be considered here. The outer 
membrane, which is so prominent just after the eggs have been discharged 
into the water (Fig. 42), is very difficult to see in sectional material. 
The second membrane is clear and since it is rather heavy its size is not 
affected greatly by preservatives. Its form is not spherical but is some- 
what longer than broad. Through the long axis it measures from 28 to 
36u, the average being about 344. The contained oncosphere measures 
rom 16 to 24u in diameter, the average being about 20u. 

The eggs of this species are larger than those of C. giganteuwm. Fuhr- 
mann reports the eggs of C. solidum as being small; ‘“‘die Oncosphaere 
hat einen Durchmesser von 0.013 to 0.016 mm, die dussere Schale einen 
solchen von 0.020 to 0.024 mm” Janicki remarks: “Die reifen Eier von 
C. solidum sind ausserordentlich klein, sie messen nur 0.020 mm im Durch- 
messer, erscheinen aber sehr charakteristisch, so dass sie nicht leicht mit 
Eiern eines anderen Cestoden verwechselt werden kénnen.”’ 

The foregoing description indicates that C. fimbriatum bears a close 
resemblance to C. solidum, but the following comparison shows some 
very outstanding differences between the two forms. 


Foramina No. testes Size of second Size of 

secundaria egg membrane oncosphere 
C. solidum Present 140 to 180 20 to 24u 13 to 16p 
C. jimbriatum Absent 100 to 125 28 to 36u 16 to 24y 


A lack of specific information on a great many points in the descrip- 
tions of C. solidum prevents a more complete comparison. The above 
data, however, are sufficient to mark the cestode under consideration 
as distinct from C. solidum. It is therefore regarded as a new species, 
which is designated as Corallobothrium fimbriatum because of the fringed 
character of the scolex. 


30 ILLINOIS BIOLOGICAL MONOGRAPHS [284 


DISTRIBUTION, ABUNDANCE AND SEASONAL 
OCCURRENCE 


Corallobothrium giganteum and C. fimbriatum have been found associ- 
ated in the intestinal tract of Ictalurus punctatus, Ameiurus melas and 
Leptops olivaris taken from the Rock, Mississippi and Illinois rivers. 
An intensive study was made only of J. punctatus from the Rock River, 
130 of that species being examined during 1926. Nearly 70 per cent 
showed infection with one or both species of Corallobothrium. The 
following table is confined to the data on the adult parasites. 


TaBLeE I 
OccURRENCE OF ADULT CORALLOBOTHRIUM 


Number Number 
Wate oct Number Stream C. fimbriatum | C. giganteum 
Examined 
Present|Average|Present|Average 
October, 1924 | A. melas 3 Rock River 0 0 0 0 
é # I. punctatus 5 € 0 0 0 
June, 1925 | I. punctatus 8 € . 16 2 21 2.6 
y “ | I. punctatus 1‘ |Mississippi R. 10 | 10 8 8 
July, 1925 | L. olivaris 1 « e 1 1 1 1 
December, 1926 | A. melas 2 Illinois River 0 0 0 0 
November, 1927 | A. melas 5 & « 0 0 0 0 
April, 1926 | I. punctatus 14 Rock River 0 0 0 0 
June, : = c 10 e 25 2:5 15 5 
July, is ie a 46 « oy 42 0.95 61 ai) 
August, 6 « ae 25 “ o 12 0.48 6 0.24 
September, “ « « 6 i « 0 0 4 0.66 
November, “ é < 11 é ss 0 0 0 0 
December, “ é “ 18 € # 0 0 0 0 


It is evident from the preceeding data that the adult form of Cor- 
allobothrium appears in the late spring or early summer, reaches its 
maximum during June and July, and disappears entirely the latter part 
of October or the first part of November. Additional data on the sea- 
sonal occurence of Corallobothrium giganteum and C. fimbriatum have been 
secured by an examination of the parasites collected by Mr. R.E. Richard- 
son of the Illinois Natural History Survey, in connection with his studies 
on the food of Ictalurus punctatus. Only the stomach and about 5 cm 
of the intestine of each fish were preserved for his investigation. From 
such a limited portion of the intestinal tract, at best, only a small per 


285] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 31 


cent of the parasites present in each fish could be secured. In all, 1252 
individuals were examined. From 954 J. punctatus, which were collected 
from June to September, 35 showed the presence of either one or both 
species of Corallobothrium. Among 278 of the same species of fish, col- 
lected from October to May, no parasites were recorded. While to be sure, 
a much larger number was examined during the period from June to 
September nevertheless a sufficient number was studied throughout the 
months from October to May to warrant the expectation of a proportionate 
percentage of parasitized fish, had the cestodes been present to the same 
degree during the whole year. Taken independently, these data would 
not be significant, but in connection with the evidence shown in Table 
1 there can remain little doubt that the adult form of both species of 
Corallobothrium occurs only from spring to fall in J. punctatus. 

Meggitt (1914) in his study of Proteocephalus filicollis states: ‘‘Almost 
every fish in autumn was infected with one or more of these parasites, 
75 per cent of which were adult; in winter, the number of infected fish 
was considerably smaller, and adults were rare; while in spring, the pro- 
portion of adults again increased.” He goes on to say that von Linstow 
failed to find adult P. filicollis at all in winter, and Zschokke noticed it only 
three times. Wagner (1917) makes the following statement regarding P. 
torulosus: ‘‘Wie verschiedene Autoren (Zschokke, v. Linstow, Kraemer, 
Riggenbach) iibereinstimmend berichten, fillt die Reife der Geschlechts- 
produkte der Fishtaénien in Zeit zwischen Friihling und Herbst. Im 
Winter sind immer nur junge, noch nicht geschlechtsreife Tiere gefunden 
worden, was auch meine Erfahrungen an /. torulosa bestatigen.” Thus 
my findings for Corallobothrium agree, for the most part, with those 
or Proteocephalus filicollis and P. torulosus as just quoted. 


32 ILLINOIS BIOLOGICAL MONOGRAPHS [286 


LIFE HISTORY OF CORALLOBOTHRIUM 


Gruber, (1878) in his study of the copepods of Lake Constance, was 
the first to discover a larval cestode in the body cavity of a Cyclops. 
He found and described a procercoid from C. brevicaudatus and made 
the following conjecture: ‘‘Die Entwicklung zur Taenia erfahrt der 
Wurm ohne Zweifel im Darme eines der zahlreichen Fische welche sich von 
den kleiner Krustern des Sees ernéhren und es méchte wohl am wahrschein- 
lichsten sein, der Jungendzustand der TJ. torulosa ist, welche nach Ru- 
dolphi und Dujardin in Cyprinoiden unser Siisswasserseen lebt, obgleich 
es mir bis jetzt noch nicht gelungen ist, dieselbe aufzufinden.” Following 
Gruber’s discovery papers appeared by Mrazek, v. Linstow, and others, 
which described procercoids from Cyclops, Diaptomus, Gammarus and 
various ostracods. In many instances the larvae were identified with 
known adult parasites, but such studies usually admit of considerable 
doubt. Schmidt (1894) was the first to feed tapeworm eggs experiment- 
ally to the smaller aquatic crustacea. He succeeeded in infecting Cypris 
ovata with the eggs of the duck tapeworm, Taenia anatina, and described 
the development of the latter species from the oncosphere to the mature 
procercoid (Cysticerkoide). 

Linton (1891) published a contribution to the life-history of Di- 
bothrium cordiceps in which he says, ‘I have found a large Dibothrium 
in the white pelican (Pelecanus erythrorhynchus) which is evidently the 
adult form of D. cordiceps, of which the trout (Salmo mykiss) is 
the intermediate host.’’ Linton gives no experimental evidence to sub- 
stantiate his supposition. Consequently his conclusions are largely con- 
jectural. No experimental work was done on the first intermediate hosts 
of fish cestodes until the work of Schneider (1903) who infected Gam- 
marus locusta with the eggs of a species of Proteocephalus, Barbieri 
(1909) described a new cestode from Alsosa finta var. lacustris which 
he called Ichthyotaenia agonis. On insufficient evidence he gives the inter- 
mediate hosts for his cestode as Bythotrephes and Leptodora. 

A successful study of the complete life-cycle of a fish cestode was 
made by Meggitt (1914), who traced the complete development of Pro- 
teocephalus filicollis by experimental methods. Cyclops varius was found 
to be the first and only intermediate host. P. filicollis is parasitic in the 
stickleback. The latter is infected by ingesting Cyclops which contain 
mature procercoids of P. filicollis. 

Wagner (1917) made a splendid experimental study of the develop- 
mental cycle of Proteocephalus torulosus, for which he discovered Cyclops 
strenuus and Diaptomus castor as the first intermediate hosts. The fish 


287] STRUCTURE AND DEVELOPMENT OF CORALLOBOTH RIUM—ESSEX 33 


host, Cyprinus orfus, is infected with P. torulosus by feeding on cope- 
pods, some of which are infected. 

Janicki and Rosen (1917) published the results of their successful 
study on the manner in which fish were infected with the plerocercoid of 
Diphyllobothrium latum. 

Following this work, Rosen (1918) elucidated the life-history of two 
other species, viz., Triaenophorus nodulosus and Abothrium infundi- 
buliforme. A year later the same worker outlined the development of 
Ligula simplicissima. It was found that the life-cycles of these forms were 
almost identical, excepting in A. infundibuliforme, the mature egg gives 
rise to a ciliated larva; the egg of that species, however, produces in the 
water an unciliated larva. In each case these larvae are eaten by some 
species of Cyclops and the subsequent development is practically the same. 
All of these cestodes except A. infundibuliforme require a second inter- 
mediate host, which is found among the young fish inhabiting the same 
waters. The adult host of D. /atum is man, the dog, or possibly the cat; 
that of T. nodulosus and A. infundibuliforme is found among the fish, 
while that of L. simplicissima is some species of aquatic bird. 

The literature contains no further studies on the life cycles of fish 
cestodes until the appearance of a fine paper by Kuczkowski (1925). 
This worker succeeded in infecting Cyclops strenuus, C. serrulatus and 
C. oithonoides experimentally with the eggs of Proteocephalus percae 
which is parasitic in Gasterosteus aculeatus, and likewise C. strenuus 
and C. serrulatus with the eggs of P. longicollis which is parasitic in Core- 
gonus albula. In this study, Kuczkowski, gave particular attention to 
the development of the bladder appendage, or ‘‘cercomer,”’ and its bear- 
ing on the “‘cercomer theory.” Bangham (1925), in his studies of the 
cestode parasites of the black bass, reports that the procercoids of P. 
pearsei were found in a species of Cyclops and also in Epischura lacustris. 
Since no experimental work was done to establish the identity of the lar- 
vae there remains considerable doubt whether they were the procer- 
coids of P. pearsei or some other Proteocephalid. 

The papers just mentioned represent the work that has been done 
on the life-cycles of fish cestodes in Europe and America up to the present 
time. Therefore the study of the developmental history of fish cestodes 
from American hosts offers a nearly unexplored field. The investigation 
reported here was undertaken in the hope that some definite information 
might be obtained on the developmental cycle of the two species of Ameri- 
can fish cestodes just described. 


OBSERVATIONS CONCERNING THE EGGS 


After the cestodes were removed from the intestine of the fish, they 
were washed immediately in tap water. This was done by grasping the 


34 ILLINOIS BIOLOGICAL MONOGRAPHS [288 


worm near the middle with a pair of forceps and rapidly raising and lower- 
ing it in the water. Each adult individual was then placed in a separate 
watchglass and covered with cold water. The excessive contractions of 
the worms caused the eggs to be ejected in milk-colored streams from 
the uterine pores of all the ripe proglottids. It was estimated that upward 
of a million eggs were emitted by an average-sized adult Corallobothrium 
gigantewm, and one-half million or more by an adult C. fimbriatum. Ac- 
cording to LaRue (1914) the eggs of the Proteocephalids usually have 
three membranes. The outermost membrane is thin, hyaline and spheroi- 
dal in form. The middle membrane is thick and granular. The inner- 
most membrane is a clear, delicate but tough structure which is closely 
applied to the embryo. 

The eggs of Corallobothrium fimbriatum are typically spherical, usually 
flattened and depressed at each pore. In their general outline they recall 
the form of an apple. The diameter varies from 0.08 to 0.14 mm. The 
outer membrane encloses a thick layer of transparent, gelatinous sub- 
stance wich is responsible for the unusually large size of these eggs (Fig. 
42). At the center of the gelatinous material is found a second membrane 
which is ovoidal in form, varying in size from 36 to 38u by 30 to 32u. 
The structure of this membrane is homogeneous and firm. It does not 
have a granular appearance in the living material nor in the eggs sec- 
tioned in the uterus. It presents rather the character of a chitinous cover- 
ing which closely resembles, and is doubless homologous to, the shell of 
the Bothriocephalan egg. Meggitt (1914) reports an aperture in the second 
membrane of the eggs of P. filicollis. I have not observed such an open- 
ing in the eggs of either species of Corallobothrium. Lying beneath the 
second membrane of the eggs of Corallobothrium fimbriatum is a dense layer 
of granular substance about 0.10 mm thick. This material is doubtless 
composed of vitelline granules and other substances, either stored for 
the nourishment of the oncosphere or cast off during its formation. Sur- 
rounded by the granular layer just mentioned is found the six-hooked 
embryo. A third membrane could not be distinguished in either living 
or sectional material. The membranes of the living egg are so trans- 
parent that the oncosphere, which is about 20y in diameter, can be seen 
distinctly. By a series of gliding movements it turns itself about within 
the shell or second membrane. Coincident with the movements of the body, 
the hooks are repeatedly extended and withdrawn. Observations made 
successively on the same individual for 12 hours showed that the onco- 
sphere continued its apparent efforts to escape from its enclosing mem- 
branes. Its movements were rather spasmodic and intermittent. A period 
of vigorous contortions, during which it would frequently turn itself 
completely about, would be followed by a longer period of inactivity. On 
several occasions oncospheres were observed to work their way through 


289] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 35 


the granular layer and reach the shell, against which they repeatedly 
brought their hooks with all possible force but without apparent effect. 
The second membrane or shell seemed impervious to the action of the 
hooks. These observations indicated clearly that the oncospheres could 
not escape from the egg membranes by their own efforts. 

Frequently in studying the eggs the rupture of the shell was noted. This 
permitted the escape of the oncosphere into the gelatinous covering (Figs. 
43, 46). I could not always attribute the ruptured shell to the pressure of 
the coverglass. Meggitt (1914) who observed the same phenomenon in 
the eggs of P. filicollis offers the following explanation: ‘The cause of its 
[the oncosphere’s] escape may be possibly due to osmosis, but it is more 
probable that it is due to the oncospheric movements.” That is, the 
oncosphere forces its way out of the shell through the aperture which has 
already been mentioned. My observations on the structure of the shell 
and the activity of the oncosphere of Corallobothrium fimbriatum make the 
latter explanation untenable. When the oncosphere is found free in the 
gelatinous covering its structure and movements are more easily observed. 
The body of the oncosphere is covered by a very delicate membrane in 
which the proximal ends of the hooks are embedded. Within this mem- 
brane is the plasma, a grayish mass of homogeneous substance containing 
extremely fine granules. The plasma vividly recalls the color and structure 
of the protoplasm found in Amoeba proteus. Cell boundaries could not be 
distinguished, and it was quite impossible to detect the presence of any 
structures that might be interpreted as muscle fibers. 

To determine the method by which the hooks of the oncosphere were 
brought into action, a long series of observations was necessary. The hooks 
are arranged in three pairs near the periphery of one pole of the embryo. 
Two pairs are placed laterally, while the third pair lies between. Usually 
the proximal ends of the hooks lie close together while the distal ends, 
bearing the hooks proper, are more widely separated. Before being thrust 
out the lateral pairs are brought up close to the middle pair. When this 
movement is completed the hooks of each pair are parallel with each other 
(Fig. 46). After watching their movements for hours, it became quite 
evident that the hooks were incapable of independent action; that their 
extension only occurred when the body of the oncosphere was elongated 
(Fig. 43); and that their return to a position of rest occurred only when the 
oncosphere again assumed a spherical form, i.e., the elongation of the 
oncosphere extended the hooks and its contraction withdrew them. The 
presence of muscle fibers attached to the hooks is extremely doubtful. 
My observations lead me to the conclusion reached by Janicki and Rosen 
(1917) in their study of Diphyllobothrium latum, viz., that the hooks, 
embedded in the membranous covering of the oncosphere, are brought into 
action as a result of the movements of the plasma rather than through the 
agency of muscle fibers. 


36 ILLINOIS BIOLOGICAL MONOGRAPHS [290 


In Corallobothrium giganteum only a glance at the eggs is needed to 
distinguish them from those of C. fimbriatum (Figs. 42, 50). The differ- 
ences in shape and size are noticed at once. They are much smaller, measur- 
ing only from 30 to 60u in diameter. Their shape is extremely irregular 
and varied. These differences are due largely to the distribution of the 
gelatinous material which surrounds the shell. It usually covers the shell 
in varying degrees of thickness, with here and there a finger-like projection 
which may give the egg a star shape. In some instances these projections 
are absent or less pronounced (Figs. 49, 50). The thickness of the gela- 
tinous material is then more uniform (Fig. 49). The shell itself is almost a 
perfect sphere, measuring from 21 to 24y in diameter, which is considerably 
smaller than that of C. fimbriatum. A layer of finely granular material is 
present within the shell, but its thickness is much less than in C. fimbriatum. 
The oncosphere, which ranges from 13 to 16u in diameter, is not sur- 
rounded by a third membrane. 

Although the structure of the eggs of the two species of Corallobothrium 
is identical, there is wide variation in the amount of the differentiated 
parts. Thus the ova of C. giganteum have less of the outer gelatinous sub- 
stance, less of the granular material within the shell, and a much smaller 
oncosphere. Also, the shape and size of the egg as a whole, and the shape 
and size of the shell are quite different in the two species. 


Experiments with the Eggs 


To determine whether or not the eggs would hatch, watchglasses 
containing several thousand eggs were placed in the dark at room tempera- 
ture. As a control another watchglass containing a like number of eggs 
was placed in the light at room temperature. Neither batch of eggs hatched. 
This experiment, together with observations on the structure of the eggs 
of each species, indicated that they did not give rise to free larvae but must 
be ingested in toto by the first host. 

It was difficult to determine the viability of the eggs because of the 
action of bacteria. Thirty-six to forty-eight hours after the isolation of 
the ova myriads of bacteria began their destructive action upon them 
but feeding experiments with eggs which had been isolated for four days 
showed positive results. Eggs liberated in nature would very probably 
remain viable for a much longer period, since the bacteria should be less 
numerous in the open waters than under the restricted conditions in the 
watchglasses. In a medium containing a minimum number of bacteria 
it is probable that the ova remain viable for eight or ten days, and possibly 
for a longer period. : 

To ascertain the manner in which the eggs were disseminated—whether 
discharged from the proglottid before or after leaving the host—a series 
of observations were made on live fish. Adult Ictalurus punctatus were 


291] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 37 


placed in the laboratory aquaria, and for three or four successive days 
after their arrival the feces were examined once or twice a day. The feces 
were obtained by placing the fish on its back and firmly stroking the ab- 
domen in the direction of the anus. The feces forced out in this way were 
inspected for the presence of cestode proglottids, eggs, etc. In such exam- 
inations of adult fish, eggs were present, but no detached proglottids were 
found. Furthermore, in post mortem examinations of over 100 adult 
I. punctatus, not a single detached ripe proglottid was recorded. From this 
evidence and that gained from the study of the adult worm (uterine pore, 
firmly attached proglottids, etc.), it is concluded that the eggs are forced 
through the uterine pores and pass out with the feces into the water, where 
they float about for a time and then sink to the bottom. 


THE FIRST INTERMEDIATE HOST 


In attacking the problem of the life-cycle of these parasites, two 
methods of approach were considered. First, it seemed reasonable to 
suppose that some clue to the intermediate hosts of the parasites might 
be gained by a thorough examination of the stomach and intestinal con- 
tents of the fish. Since the larvae of the cestodes probably must enter their 
final host passively with its food, a systematic study of the food of the fish 
was made a part of the routine of examinations. Secondly, the inter- 
mediate host might be discovered through direct experiment, i.e., by 
feeding ripe proglottids or eggs to any invertebrate animals which might 
possibly serve as the first intermediate host of the parasites. 

The diet of 55 Ictalurus punctatus examined during the months of June 
and July consisted almost exclusively of crayfish, Cambarus propinquus, 
larval and adult Hexagenia bilineata, other insect larvae, and portions of 
mollusks and fish. The first two of these forms were considered as possible 
intermediate hosts, and macroscopical and microscopical examinations 
were made of the remains of those obtained from the stomachs of the fish, 
but with negative results. It was thought advisable, however, to attempt 
an infection by feeding them ripe proglottids or eggs. 

Experiments with the crayfish, Cambarus propinquus, were carried on 
from July fifth to fourteenth. Crayfish secured from Rock River were 
placed in laboratory aquaria. After they had been allowed to fast for two 
or three days, living adult individuals of C. giganteum and C. fimbriaium 
were fed to them. Although they did not show a decided preference for 
the worms, they did eat them. At intervals varying from a few hours to a 
week after the worms had been ingested, the crayfish were examined. The 
intestine was removed, its contents teased out on a slide and inspected 
for the presence of oncospheres that might be in the lumen or for more 
advanced stages of the larvae encysted in the intestinal wall. Likewise, 
all the organs contained in the body-cavity were examined, and also the 
surrounding musculature. The results were negative. 


38 ILLINOIS BIOLOGICAL MONOGRAPHS (292 


Experiments with the Mayfly, Hexagenia bilineata, were made between 
July twentieth and August fifteenth. While the work with the crayfish 
was in progress, experiments were conducted on the larvae of Hexagenia 
bilineata. To obtain uninfected material adults were caught and stripped 
of their eggs, which were placed in large Petri dishes in the laboratory where 
they were allowed to stand. The water on them was changed every 24 
to 48 hours. Seventeen days incubation produced hundreds of small larvae. 
Eggs of C. giganteum and C. fimbriatum were placed in two Petri dishes 
containing fifteen to twenty of these larvae and water-plant. Observations 
of the larvae under the binocular showed them swimming about or clinging 
to the vegetation in the containers. None of them seemed to be attracted 
to the tapeworm eggs. Subsequent examination of the larvae showed no 
trace of infection with the cestode larvae. 

Experiments with Cyclops albidus also were conducted during July; 
these were coincident with the experiments just described. A supply of 
Cyclops albidus was collected from a lagoon near Rock River. A single 
individual was placed in each of six watchglasses, along with a spray of 
water-plant. After three or four days it was decided that they could live 
under such restricted conditions, as all of them were alive and very active. 
Then a drop of water containing the eggs of Corallobothrium giganteum 
was added to each of three watchglasses; and to each of the other three, 
a drop of water containing the eggs of C. fimbriatum. My observations of 
the Cyclops under the binocular convinced me that they were not attracted 
to the cestode eggs. They would, however, readily devour protozoa which 
were placed with them. On one occasion fork-tailed cercaria, obtained 
from a snail of the genus Physa, were placed in a watchglass with a C. 
albidus. To my great astonishment the Cyclops pounced upon one after 
another until fourteen were devoured.* Since efforts to observe the in- 
gestion of the eggs by this species of Cyclops resulted negatively, and 
since no oncospheres or larvae were found when all of the individuals were 
afterward examined, it was concluded that further experiments with this 
species would be futile. It was evident that such a process of elimination 
applied to each animal which might be suspected of being an intermediate 
host of these cestodes, would consume much more time than I had at my 
disposal. Therefore, a more comprehensive method was conceived and 
pursued. 

Mass infection of plankton was accordingly tried from July twentieth 
to August second. Not far from the Rockford College campus Rock River 
is obstructed by a dam. By holding a plankton net in the water that poured 
over this dam, it was possible in a few minutes to collect samples of a large 
number of the pelagic forms, as well as some bottom organisms that were 


* This may account for the infrequency with which cercaria are met in plankton samples. 


293] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 39 


caught in the current and carried over the dam. A large quantity of the 
material gathered by this means was placed in each of two crystalizing 
dishes. Water-plant of the genus Cladophora taken from the face of the 
dam was placed in each dish. The bottom of one dish was strewn with the 
eggs of Corallobothrium giganteum and that of the other with the eggs of 
C. fimbriatum. Under these conditions all species were subjected to identi- 
cal conditions; each was given an equal opportunity to ingest the eggs. 
It was hoped by an examination of the species contained in these cultures 
that the primary host of each worm might be discovered. After the 
cultures had stood for eight days, an examination of the different species 
present was begun. Since among the Copepoda a number of species had 
been discovered as the primary hosts of cestodes, representatives of that 
group were examined first. Each copepod was placed on a slide in a small 
drop of water, then excess water was drawn off with a fine pointed pipette 
so that only enough water was left to cover the specimen. This procedure 
minimized its movements and anchored it to the spot so that it could be 
examined successfully under the microscope. By this method several 
hundred copepods were examined. 

From the culture in which C. giganteum eggs were placed, Cyclops 
serrulatus were found to be infected with from 1 to 6 larvae each, while 
C. prasinus were infected with from 1 to 3 each. From the culture inocu- 
lated with C. fimbriatum eggs, infected individuals were found among 
C. bicuspidatus and C. serrulatus, the former usually showing much heavier 
infection. No infection was observed among the copepods, C. albidus, 
C. fuscus, C. bicolor, or Diaptomus, nor among the Cladocera examined. 

Since these results were accepted as a clear indication of the true first 
intermediate hosts of Corallobothrium giganteum and C. fimbriatum, further 
group infections were discontinued and a series of experiments were carried 
on with isolated groups of the species that had shown infection under mass 
conditions. 

To determine the species and to prevent the inclusion in the experi- 
mental groups of individuals infected in nature each Cyclops was examined 
microscopically. Because of the transparency of the Cyclops, a cestode 
larva, when present, could be detected without great difficulty. The un- 
infected C. serrulatus and C. prasinus were placed together, and uninfected 
C. bicuspidatus and C. serrulatus were also allowed to occupy the same 
container. 

Experimental infection of Cyclops was successfully attempted in 
August. These copepods were placed in a fingerbowl of tap water to which 
were added a few sprays of water-plant that had previously been rinsed 
thoroughly to reduce the protozoa in the fingerbowls as much as possible 
and likewise to prevent the entrance of uninspected Cyclops. On August 
twentieth at 11:30 a.m. the eggs of C. fimbriatum were placed in with 


40 ILLINOIS BIOLOGICAL MONOGRAPHS [294 


them. Oncospheres were found at 3:45 p.m. of the same day in the body- 
cavity of the Cyclops examined. Thus in a little more than four hours the 
oncospheres had migrated from the intestine into the body-cavity. 

To determine whether or not the ingestion of the tapeworm eggs was 
selective or accidental, a study of the feeding habits of the Cyclops was 
andertaken. A fingerbowl containing C. bicuspidatus and C. serrulatus and 
the eggs of Corallobothrium fimbriatum was observed under the binocular. 
It was noted that the Cyclops foraged over the sprays of water-plant in 
the bottom of the fingerbowl and browsed on the particles of debris ad- 
hering to the vegetation and to the bottom. Any of the smaller protozoa 
which came near were instantly consumed. The outer gelatinous portion 
of the tapeworm eggs was repeatedly trimmed off and eaten, while the 
inner membrane (shell) containing the oncosphere was rejected. By con- 
tinuous observation it was revealed that occasionally the inner portion 
also was taken in with the rest, which indicated that the eggs were eaten 
only incidentally, along with other organic material on which the Cyclops 
fed. This condition is in striking contrast with that of Diphyllobothrium 
latum and related forms in which the eggs give rise to ciliated larvae that 
successfully simulate protozoa and therefore constitute real objects of 
prey, attracting the Cyclops by their movements and tempting them to 
pursuit and capture. 

The mass infection experiments had shown that Cyclops serrulatus 
could be infected by feeding the eggs of either species of Corallobothrium, 
but this point was not settled in respect to Cyclops bicuspidatus and 
C. prasinus. Therefore the eggs of Corallobothvium giganteum were fed to 
Cyclops bicuspidatus and those of Corallobothrium fimbriatum to Cyclops 
prasinus. The eggs of Corallobothrium giganteum were eaten by Cyclops 

, bicuspidatus and the oncospheres migrated to the body-cavity, but no 
development was observed. The oncospheres, five days after the eggs were 
fed, measured only 16u in diameter, their original size. The number present 
ranged from 2 to 15. This condition was noted in 15 different individuals 
examined from 1 to 5 days after feeding. The infection of C. prasinus with 
Corallobothrium fimbriatum larvae was light; only 2 individuals out of 10 
harbored the larvae. The latter, however, were well developed. 


Development of Corallobothrium fimbriatum in Cyclops 


When Cyclops bicuspidatus and C. serrulatus had been identified as the 
first intermediate hosts of Corallobothrium fimbriatum, a study of the 
successive developmental stages in these copepods was begun. All observa- 
tions were made on living material and within the period from August 
twentieth to September fourth. Beginning from one to four hours after 
feeding, the progress of development was traced through all of the stages 
found in these animals. As has already been stated, oncospheres were 


295] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 41 


first observed in the body-cavity of the Cyclops about four hours after 
the eggs had been placed in the watchglasses. Immediately following their 
liberation in the intestine of the Cyclops, the oncospheres increased 
slightly in diameter. While contained in the egg membranes, they measured 
from 16 to 20y, but after liberation they measured from 18 to 23 in 
diameter. Those present in the body-cavity about four hours after the 
eggs were fed measured from 20 to 25u. Their form, however, constantly 
changed. They did not remain fixed to the intestinal wall of the host by 
means of their hooks but kept up almost constant motion. Since the 
elongation and contraction of the body put the hooks in operation, the 
action of the oncospheres resulted in the repeated extension and with- 
drawal of their hooks. 

After the oncosphere had gained the body-cavity of the Cyclops, 
development proceeded very rapidly. Twenty-four hours after the eggs 
had been fed, four oncospheres were liberated, these measured from 30 
to 40u according to the stage of contraction. Figure 53 shows one as it 
appeared within the abdomen of the Cyclops, while figures 44 and 45 
represent their condition 5 to 10 minutes after being liberated. Clear spheri- 
cal bodies (cells) surrounded by heavy granules were seen throughout the 
body mass. When first liberated the oncospheres moved about with a 
gliding movement. During this time the hooks were kept in almost con- 
stant motion. Five or ten minutes after the larvae had been removed from 
the Cyclops indications of degeneration were seen. Then a transparent 
membrane was pushed out from the body of the oncosphere (Figs. 44, 45). 
In the individual represented in figure 45 there was a secondary membrane 
within the outer one from which the parenchyma had withdrawn. Thus it 
appears that the larvae at this stage possess two membranes; the outer 
may be considered cuticular while the inner may represent the basement 
membrane. 

Seventy-two hours after feeding, the larvae ranged from 25 to 70y in 
diameter (Figs. 55, 48). They were capable of distinct contractions which 
could be witnessed while they were still within the host. Upon removal, 
which was accomplished by tearing the Cyclops open by means of two 
sharply pointed needles, they appeared as a thin-walled sac containing 
globular bodies representing the parenchymatous tissue, which resembled 
an emulsion. Their movements were extremely feeble and continued for 
only a few minutes, then a spherical form was assumed and degeneration 
soon followed. The hooks, which were widely separated at one pole of the 
body at this stage, were incapable of effective action. 

Four days after feeding, 12 oncospheres, measuring from 25y in diameter 
to 135y in length by 60u in width, were removed from the body-cavity of 
one Cyclops. Figure 54 represents the most highly developed individual. 
Its sac-like body showed considerable differentiation. A cuticular mem- 


42 ILLINOIS BIOLOGICAL MONOGRAPHS [296 


brane surrounded the body and beneath it the tissue had a striated appear- 
ance, due to the development of the subcuticular musculature. Large cells 
surrounded by more or less heavy granules were evident. A few small 
calcareous bodies were to be seen scattered here and there throughout the 
body mass. 

On the fifth day a Cyclops was dissected and 14 larvae were present 
in the body-cavity. These were from 0.06 to 0.24 mm in length. Except for 
the increase in size, no marked difference between these and the four-day 
individuals was noted. 

One Cyclops observed on the sixth day had 16 larvae in the body-cavity 
(Fig. 51). Some of these larvae showed an increase in size over those of the 
fifth day, since they measured from 0.22 to 0.4 mm in length, according 
to the state of contraction. Upon being liberated, they moved about more 
vigorously for several minutes. The narrower end, which bore the hooks, 
showed more activity than the broader portion. Further differentiation 
had occurred, as the pole at which the hooks appeared presented a striking 
contrast to the homogeneous structure of the remainder of the body. It 
had become extremely transparent. The granules which appeared in the 
remainder of the body were entirely absent in this portion. At the proximal 
limit of this transparent region, or about 0.04 mm from the extreme body 
limit, a slight constriction appeared. In some individuals the hooks were 
attached to this portion, but in others they occurred just beyond the 
constriction in the granular region of the body (Fig. 56). In a number of 
cases some of the hooks were present on both regions (Fig. 57). 

After six days no further increase in the length of the larvae took place, 
but many changes in their structural aspects occurred in quick succession. 
Thus on the seventh day there were evidences of differentiation at the pole 
opposite that on which the hooks were found. On the eighth and ninth days 
the larvae showed three distinct regions marked by two constrictions; 
one delimited the transparent region, the other occurred about one-third 
of the body length from the pole opposite that which bore the hooks 
(Fig. 56). By the tenth day, outlines of the developing suckers could be 
seen in this portion of the body, which was, therefore, the potential scolex. 

Larvae studied on the eleventh day after the Cyclops were fed showed 
the four suckers well developed and at the anterior extremity a distinct 
end-organ. The large globular cells of the scolex region had disappeared. 
In their place a reticulum of minute, rounded cells with an occasional 
muscle fiber had appeared. The calcareous bodies, 5 or 6 in number, were 
confined, with a very few exceptions, to the middle portion of the larva 
The transparent region, more distinctly separated from the middle portion, 
formed a bladder-like appendage or cercomer (Fig. 57). 


297] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 43 


The development on the twelfth and thirteenth days differed from that 
on the eleventh in the following respects: The scolex, which had been 
differentiated by the eleventh day, was invaginated; the number of cal- 
careous bodies had increased; and the length, by reason of the invaginated 
scolex, was somewhat reduced (Fig. 58). 

Development in the Cyclops is completed by the fourteenth or fifteenth 
day after the ingestion of the eggs. The best indication of this is the 
invagination of the scolex and the loss of the bladder, or posterior ap- 
pendage. Janicki and Rosen (1917) have given this larva the name of 
procercoid. After this stage is reached the parasite must enter another host 
before further development can be attained. 

To indicate the extent of infection that may occur in such experimgnts, 
it should be stated here that from one culture 50 specimens of C. bicuspi 
daius were removed and 46 contained the larva of Corallobothrium fim- 
briatum. 


Development of Corallobothrium giganteum ix Cyclops 


The development of this species corresponds closely with that of 
Corallobothrium jfimbriatum. The oncospheres were observed in the body- 
cavity of Cyclops serrulatus eight hours after the eggs had been placed in 
the fingerbowls. Their presence was more difficult to detect because of 
their smaller size and greater transparency (Fig. 52). The number of 
larvae present in these species of Cyclops varied from 1 to 8. Therefore 
the infection was less intense than that found in the previous species where 
as many as 18 larvae were present. 

No further observations on the progress of this species were recorded 
until the eighth day after the eggs were fed. By that time the larvae had 
attained a length of from 0.28 to 0.39 mm, which corresponded closely to 
the size of the larvae of Corallobothrium fimbriatum at the end of an equal 
period of time. However, the differentiation of various body regions had 
progressed considerably more than was true of the C. fimbriatum larvae 
in the same length of time. The suckers and end-organ were distinct and 
the bladder, or posterior appendage, was almost completely separated 
from the middle portion; and a larger number of calcareous bodies were 
present in this species than occurred in C. fimbriatum at a much later 
period. It was also noted that the globular cells, which were very prominent 
in C. fimbriatum, were much less pronounced in these larvae. Aside from 
the heavy granules scattered through the body, the structure was more 
homogeneous than that of the other species (Fig. 70). 

On the twelfth day an examination of infected Cyclops revealed that 
the larvae had already become mature procercoids (Fig. 68). This was at 
least two days earlier than the same stage was reached by C. fimbriatum. 
The cuticula had increased greatly in thickness. The length of the larvae, 


44 ILLINOIS BIOLOGICAL MONOGRAPHS [298 


which was somewhat reduced by the invagination of the scolex, ranged from 
0.2 to 0.25 mm according to the degree of contraction. As was true in the 
case of the other species, the bladder or posterior appendage had been 
shed and in addition a well-developed excretory vesicle had appeared in 
the posterior portion of the body. This organ, which measured from 5 to 9u 
in diameter, could be traced anteriorly for about 0.06 mm when it became 
obscured by the surrounding structures. Efforts to detect smaller vessels, 
which doubtless emptied into it, failed entirely. The vesicle terminated 
posteriorly in a duct which emptied at the point where the bladder had 
been attached. Pulsations which began at the anterior portion and passed 
to the excretory pore, could be seen to proceed rhythmically along its 
length. An attempt to discover the vesicle in individuals that still possessed 
the bladder proved futile. There may be a relation between these two 
structures, besides one of sequence in development, but any statement 
further than this would be purely conjectural. Such a vesicle was not 
observed in C. fimbriatum. If present, it was obscured by other structures. 
It was easily seen in a later stage to be described presently. 

Twelve-day larvae possessed a greater number of calcareous bodies, 
from 15 to 25. These were not restricted as in C. fimbriatum almost ex- 
clusively to the middle body region but were distributed everywhere along 
the periphery. More of them, however, were present in the posterior 
portion than elsewhere. The total number in this species is almost twice 
that discovered in any of the larvae of C. fimbriatum of about the same age 
(Figs. 58, 68). 

The suckers and end-organ of the invaginated scolex could not be dis- 
tinguished clearly in this species, as they were obscured by the overlying 
tissues. Following their liberation from the Cyclops, the larvae moved 
about actively for an hour or more. In one or two instances the scolex was 
evaginated. Figure 64 (a to /) represents the successive movements and 
shapes assumed by the larva before the evagination of the scolex. Figure 66 
represents the appearance of one specimen with scolex everted after having 
been fixed and mounted. Maturity in the Cyclops has been reached by this 
species when the scolex has become invaginated, when the bladder appen- 
dage has been shed, and when the excretory vesicle has appeared. 

It is interesting to note the difference in the rate of development found 
in these two species. Although Corallobothrium giganteum begins with a 
much smaller oncosphere, it develops at a rate sufficient to attain maturity 
in the Cyclops about two days before that point in development is reached 
by C. fimbriatum. Furthermore, the size of the adults in the two species 
would indicate that this difference manifests itself throughout the develop- 
mental cycle of each. 

An interesting observation, which was the cause of considerable concern 
early in the course of these experiments, was made in connection with the 


299] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 45 


Cyclops that contained from 8 to 18 larvae. In such individuals several 
stages of development were frequently represented. For example, fifteen 
days after the eggs had been fed, mature procercoids and others, represent- 
ing from three to ten days progress, were present in the same Cyclops. 
These differences were first considered to be due to the variability in the 
time at which the eggs had been ingested. The mature forms would thus 
represent the ingestion of the first eggs, and the less mature individuals 
the eggs eaten more recently. Further observation, however, indicated 
that this explanation was incorrect, since it was noted, by examination 
of the same Cyclops on successive days, that certain of the larvae increased 
very little in size. Among six C. fimbriatum larvae present in one Cyclops 
serrulatus, there were two fully mature, one represented the stage shown in 
figure 57, and three showed a development of about 10 days. In another 
Cyclops of the same species containing eight larvae of C. gigantewm, three 
were mature (Fig. 68) and five were immature. Their movements after 
liberation were very feeble and lasted for only five or six minutes. One 
Cyclops bicuspidatus infected with 18 larvae of Corallobothrium fimbriatum 
showed nearly all the stages from the oncosphere to that represented in 
figure 56. This individual was examined just six days after the eggs were 
fed (Fig. 51). Many of the Cyclops examined from 25 to 30 days after the 
feeding of the eggs, showed this same condition of the larvae. This in- 
hibition of growth among the larvae was probably due to the inability of 
the Cyclops to furnish sufficient nourishment for the complete develop- 
ment of more than three or four individuals. 

In connection with the study of the development of C. giganteum an 
observation was made that caused considerable perplexity. In the body- 
cavity of several Cyclops, along with mature procercoids (Fig. 67), there 
frequently occurred as many or more very transparent, weakly contractile 
individuals devoid of hooks and calcareous bodies. They usually measured 
about 0.10 mm by 0.05 mm (Fig. 63), though one individual measured 
0.30 mm by 0.10 mm. This individual was constricted near the middle 
and presented the appearance shown in figure 61. Kuczkowski (1925) in 
his study of I. percae reports the observation of forms similar to those I 
have encountered in connection with C. gigantewm. He offers the suggestion 
that they represent the cast-off bladders (cercomers) of the mature pro- 
cercoids. JI am inclined to the same conclusion since the size of these 
transparent individuals is usually about the same as the bladder appendage. 
Furthermore, the structure of the two is almost identical. In a few in- 
stances, however, these peculiar forms were more than twice the size of 
the attached bladder (Fig. 61). Consequently, if they are accepted as 
being the bladders that have been separated from the mature procercoids, 
there must be claimed for them a certain power of independent growth. 


46 ILLINOIS BIOLOGICAL MONOGRAPHS [300 


A shifting of polarity was noted in the course of the development of 
the larvae of both species of Corallobothrium. During their early phases 
(from the oncosphere to about ten days), when removed from the Cyclops 
they moved slowly about with the end bearing the hooks directed an- 
teriorly. Coincident with the differentiation of the scolex at the opposite 
pole, their polarity was reversed and movements thereafter were made 
with the scolex-end in advance. This phenomenon was noted also by 
Janicki and Rosen (1917) in their study of Diphyllobothrium latum. 

The effects of infection on the Cyclops deserve brief consideration. 
Infection with a large number of oncospheres (20 to 50) appeared to cause 
the Cyclops no discomfort. Individuals so infected were quite as active 
as those unparasitized. Cyclops whose body-cavities contained from 1 to 
6 larvae in advanced stages of development disclosed little evidence of any 
serious effects. When the infection went beyond this point, however, the 
Cyclops showed evidences of being greatly inconvenienced by the presence 
of the larvae. An infection as intense as is shown in figure 51 almost totally 
incapacitated the Cyclops. Such individuals no longer attempted to swim 
about but settled to the bottom of the fingerbowls and only moved when 
stimulated with the point of a needle or by vigorous stirring of the water. 
Then only a few strokes were made with the antennae and abdomen, after 
which they again settled to the bottom where they remained, apparently 
dispossessed of every desire for food or action. As has been pointed out by 
Meggitt (1914) and Janicki and Rosen (1917), such individuals fall an 
easy prey to the fish which feed on them. However, according to my ob- 
servations no Cyclops that had been infected in nature contained more than 
a single procercoid. When it is considered that the chances of a multiple 
infection are rather slight in a large lake or river, the effect of the parasites 
on the Cyclops is of relatively little significance. 


THE SECOND INTERMEDIATE HOST 


The experiments in this line were carried on from August 24 to Sep- 
tember 9, 1926. Studies on the food of Ictalurus punctatus, Ameturus melas 
and Leptops olivaris had shown that minnows and occasionally other 
species of fish constituted a part of their diet. Copepods were never found 
in the food examinations of any of the foregoing species. Therefore it 
seemed improbable that such a high percentage of infection (nearly 70% 
in the case of J. punctatus) could have resulted from the accidental ingestion 
of infected Cyclops. Consequently no experiments were made on the 
direct infection of J. punctatus through the agency of the Cyclops. I in- 
clined to the belief that infection of the catfish very likely occurred through 
the ingestion of forms that fed largely on the Entomostraca of the river. 
Since the Entamostraca were known to comprise a high percentage of the 
food of minnows, it was evident that some species of minnow might act 


301] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 47 


as a second intermediate host for the species of Corallobothrium. One of 
the most widely distributed minnows (Notropis blennius) was selected as 
the first subject. These fish could not be obtained except from the open 
waters of the streams in the vicinity of the laboratory. About 100 were 
taken from Rock River on August 24 and placed in the laboratory aquaria, 
where they remained 12 days before infection experiments were begun. 
During that time the weaker individuals were eliminated by death and the 
stronger were left for the experiments. No food was given them except 
bits of bread and cracker crumbs, on which they seemed to thrive very 
well. 

From among these fish 8 individuals were examined but no larval 
cestodes were found. Although the findings which resulted from the 
examination of eight minnows are not considered as final evidence on the 
presence or absence of larvae in the remainder of the group, they may be 
taken as a fair index of the degree of infection in that particular school of 
minnows. 

From a culture containing Cyclops bicuspidatus 30 to 40 specimens in- 
fected with procercoids were removed. These were placed in an aquarium 
(2’ long, 10” wide, 12” deep) containing water about three inches deep. 
Three of the smaller minnows were selected and placed in the same aquar- 
ium on September fourth. Three days later one minnow was examined 
and three cestode larvae were discovered on the mesentery along the outer 
wall of the intestine. Very probably there were others that escaped detec- 
tion because of their extremely small size as they were only 0.2 to 0.4 mm 
in length, according to the exact degree of contraction (Fig. 60). The two 
remaining minnows were killed September ninth and preserved in formol 
for study by the section method. 

Another group of three minnows was infected in the manner just de- 
scribed on September second. On the day of my departure from Rockford, 
September ninth, these minnows were placed in a quart jar containing 
3 inches of water in which they remained for about 12 days. One of them 
died while I was enroute to Urbana. Of the two that remained one was 
killed October first, but there were no free larvae observed in the body- 
cavity. Any that may have been present had probably migrated to the 
musculature of the body-wall. The third individuei was placed in an 
aquarium with a small uninfected Ameiurus melas. The aquarium over- 
flowed shortly afterwards, and this gave the minnow an opportunity to 
escape. An examination of the catfish on April twenty-ninth gave negative 
results. 

Sections prepared from a minnow killed three days after infection 
showed two larvae in the intestine, one outside of the intestine in the body- 
cavity, one in the ovary and three in the musculature. These larvae were 
not encysted (Figs. 65, 69). Since the larvae were found alive in the body- 


48 ILLINOIS BIOLOGICAL MONOGRAPHS [302 


cavity of the minnows and since a sectioned minnow showed their presence 
in the lumen of the intestine, in the coelomic cavity near the intestine and 
in the ovary and musculature of the body-wall, it is concluded that this 
minnow (Notropis blennius), and probably others, may serve as a second 
intermediate host for Corallobothrium fimbriatum. No experiments other 
than those already described were made on the life-cycle of C. giganteum. 
However, it is very probable that the developmental cycle of this species 
closely approximates that of C. fimbriatum. 

After the conclusion of the experiments just described, I had an oppor- 
tunity to secure additional data on the life-cycle of Corallobothrium jfim- 
briatum. By the use of a specially devised trawl, Dr. David H. Thompson 
has been able to take small fish which are rarely obtained by the use of 
ordinary apparatus. In the early part of September, 1927, he secured 32 
Ictalurus punctatus and shipped them to me alive. These fish measured 
from 2 to 3 inches in length. Six individuals out of the shipment harbored 
from 1 to 3 sexually mature Corallobothrium fimbriatum. The food of the 
young I. punctatus consists of plankton, insects, and various kinds of debris. 
It is quite improbable that infection with this cestode occurred in any other 
way than by the ingestion of copepods that harbored the procercoids. 
Therefore it was evident from my studies that the infection of catfish 
with Corallobothrium fibriatum might result either by feeding on minnows 
which harbored the larvae, or by the ingestion of infected copepods. The 
evidence for the latter contention seemed quite substantial but further 
experimentation was necessary to establish the former statement. 

To determine whether or not infection with Corallobothrium fimbriatum 
might take place through the agency of a minnow, the following experiment 
was carried out. From the cestodes obtained from the small Jctalurus 
punctatus a large number of eggs were collected. These eggs were fed 
October third and fourth to uninfected Cyclops prasinus, and the pro- 
cercoids developed in about 2 weeks. On October nineteenth the Cyclops 
were placed in an aquarium with two minnows (Hybopsis storerianus)* and 
I witnessed the ingestion of the Cyclops by the minnows. Two Ameiurus 
melas which had been kept in a laboratory aquarium since November 5, 
1926, were used for the next phase of the experiment. No food had been 
given to these fish from June to September, 1927. Any parasites that they 
may have had when brought into the laboratory, it seemed reasonable to 
suppose had disappeared during their confinement. One of them was 
examined in September and no parasites of any kind were found. On 
October twenty-sixth the two minnows that had been fed the infected 
Cyclops were placed in the aquarium with the other A. melas. Both 
minnows were eaten. An examination of the catfish was made November 
eleventh and 6 larvae were recovered from the intestinal tract. They 


* Identified by Dr. G. K. Noble, American Museum of Natural History. 


303] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 49 


measured about 0.3 mm in length. When the scolex was evaginated they 
resembled very closely the larva shown in figure 64m, but when the scolex 
was invaginated they resembled in every respect the procercoid of Corallo- 
bothrium fimbriatum; so much so that I have no doubt whatever as to their 
identity. The results of this investigation make possible an explanation 
of how adult catfish, which do not feed on plankton to any appreciable 
extent, become infected with Corallobothrium. 


Plerocercoid Larvae 


As has already been stated, the completion of the growth of the larvae 
in the Cyclops has been designated as the procercoid stage. Upon entering 
the next host transformation into the plerocercoid stage is begun. In 
Corallobothrium the first step in this process is the evagination of the 
scolex. Should the Cyclops infected with the procercoid be ingested by a 
catfish, development into the plerocercoid begins at once. However, 
should the procercoid be taken into another fish such as a minnow, which 
might be considered an accommodation host, the larva migrates from the 
intestine and takes refuge in the body-cavity or some organ of the second 
host where it remains until ingested by the definitive host. Little develop- 
ment occurs before it reaches the proper fish. There was close similarity 
between the larvae recovered from the minnow and the procercoid of 
Corallobothrium fimbriatum taken from the Cyclops. The only differences 
noted were the appearance of a larger number of calcareous bodies and an 
excretory bladder which was not observed in any of the procercoids when 
removed from the Cyclops (Fig. 60). The smallest plerocercoid found in 
the intestine of the catfish measured about 0.2 mm in length. In this in- 
dividual, as in all the young plerocercoids, the scolex is the most prominent 
feature. The body is only a small cone-shaped projection in which no in- 
ternal organs can be distinguished. Not until the larvae reach a length 
of about 1 mm is there a distinct resemblance to the adult individual 
(Fig. 62). 

As has already been shown by Table I, no adult parasites belonging to 
the genus Corallobothrium were present among the fish examined from 
October to April. This was taken as decisive evidence that the adults do 
not appear before late spring or early summer and disappear entirely in 
the late fall or early winter. While this study was in progiess it was dis- 
covered that the fate of the larvae was entirely different. Because of the 
difficulty of determining accurately the species of the smaller plerocercoids 
(Fig. 59), no attempt was made to record them according to species. It 
was found, however, that larvae of both species of Corallobothrium were 
present throughout the year. Therefore in the data recorded below, the 
total number of larvae present in each fish is recorded without regard to 
the species. In the examination of 130 Ictalurus punctatus, cestodes belong- 


50 ILLINOIS BIOLOGICAL MONOGRAPHS [304 


ing to any other genus than Corallobothrium were encountered only four 
times. In those instances the parasite belonged to the genus Proteo- 
cephalus. Since the larval stages of this form would be exceedingly rare, 
they should not affect very materially the data on the plerocercoids of 
Corallobothrium. 


Taste II 


SEASONAL OCCURRENCE OF CORALLOBOTHRIUM PLEROCERCOIDS 
Data from Ictalurus punctatus taken in Rock River 


Number Number Average 
Date Examined Plerocercoids per fish 
April, 1926 14 6 0.43 
June, ¢ 10 53 Byes) 
July, 46 150 one, 
August, « 25 18 0.72 
September, “ 6 2 0233 
November, “ 11 23 2-0 
December, “ 18 49 3.3 
Total 130 301 2:3 


These data indicate that the larvae which enter the fish host in the fall 
remain in the intestine through the winter. Since those found during 
December and April measured only from 0.5 to 0.8 mm in length, it is 
evident that little growth takes place during the winter and early spring. 
This cessation of growth may be due to temperature conditions or to the 
absence of sufficient nourishment for further development. Unpublished 
data collected by the Hlinois Natural History Survey indicate that [ctalurus 
punctatus feeds very little during the winter. Doubtless both temperature 
and the lack of food are responsible for the inhibition of growth during this 
season. 


305] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 51 


COMPARISON OF PROTEOCEPHALUS AND 
CORALLOBOTHRIUM 


With the exception of the work of Rosen, all of the experimental work 
on life-histories of fish cestodes has been on species of Proteocephalus 
(=Ichthyotaenia). Megitt’s description of the development of Proteo- 
cephalus filicollis in Cyclops varius is incomplete in some respects. Briefly 
stated, he made the following observations: After the ingestion of eggs the 
oncosphere attaches itself to the wall of the alimentary canal where it 
remains about a week before breaking through into the body-cavity. After 
gaining the coelom it takes up a position near the anterior end of the 
carapace or in the head above the eye. The embryonic hooks gradually 
disappear. The body becomes covered with highly refractive bodies. The 
procercoid reached maturity at the end of three weeks. He states that the 
procercoid is an elongate gray body of variable size, without scolex or neck. 
The larva is devoid of divisions. Four suckers occur at the anterior end but. 
an apical sucker is absent. The end bearing the suckers is never invaginated 
during development. No signs of excretory organs were seen. 

The development of Proteocephalus torulosus in Cyclops strenuus and 
Diaptomus castor as described by Wagner is as follows: The oncospheres 
were observed in the coelom 24 hours after the eggs had been placed in 
with the copepods. At the end of 48 to 72 hours the larvae measured 20 
to 30u. They had settled themselves on or near the intestine. After a short 
time the larvae began to elongate. At one pole the first trace of the suckers 
appeared. Circular and longitudinal muscle fibers appeared along with 
excretory ducts and large calcareous bodies. An invagination occurred 
at the posterior end which was eventually taken over by the excretory 
bladder. The procercoid attained a length of 0.5 to 1 mm. It possessed 
four suckers whose radial muscles were distinct. The scolex possessed 
neither hooks nor rostellum. It was not set off from the rest of the body. 
The body was extremely contractile. The cuticula is broken up in a cover 
of fine hairs and bristles. In some individuals the embryonic hooks were 
strewn over the posterior half of the body. The excretory system was 
plainly visible in its entirety and consisted of an excretory bladder and 
a dorsal and ventral vessel on each side. Wagner makes no mention of a 
bladder appendage or cercomer. 

Kuczkowski’s description of the development of Proteocephalus percae 
in Cyclops strenuus may next be summarized. In the course of a few 
days after the infection, the spherical larvae of a diameter of 35u, were 
observed in the coelom of the Cyclops. This larva soon began to grow 


52 ILLINOIS BIOLOGICAL MONOGRAPHS [306 


and lose its spherical form. A vaculated region appeared within the 
body mass which the author interpreted as the “lacuna primitiva”’ of 
Grassi and Rovelli. At this stage the larva was 41 to 55u in diameter. 
There was a decided growth in length and an increased contractility. 
A cercomer appeared from 14 to 21 days after the infection, then the length 
of the larva was from 138 to 3274 and the length of the bladder from 
13 to 42u. The bladder was without hooks. These were found on the pos- 
terior portion of the larval body. The procercoid possessed four elliptical 
suckers. No apical sucker was present but a bladder-like protrusion was 
noted at the apex of the scolex. The figures indicate that the cuticula 
was smooth. A well developed excretory system was present, also cir- 
cular and longitudinal muscle fibers. The larva showed the typical cal- 
careous bodies. 

A comparison of the developmental phases of the species of Pro- 
teocephalus just described with those of Corallobothrium brings out 
some rather significant differences. In Proteocephalus the bladder ap- 
pendage or cercomer is entirely wanting in two species according to the 
descriptions given and rather rudimentary in the third. In Coralloboth- 
rium the cercomer is a decidedly prominent structure whose length equals 
one-third that of the body, 40 to 60u, and is never rudimentary. In 
Proteocephalus percae where the bladder has been observed, it shows 
signs of being rudimentary since in some cases it measures only 13y in 
length. According to Kuczkowski the cercomer of P. percae never bears © 
the embryonic hooks. These hooks are always found on the posterior 
portion of the body. This condition is likewise true of the other species 
of Proteocephalus which have been described. In Corallobothrium, how- 
ever, they may be confined to the cercomer, divided between the cercomer 
and the body, or all on the body. In Proteocephalus the suckers are 
differentiated near the anterior end of the larval body without the scolex 
being set off. The scolex is never completely invaginated. In Corallo- 
bothrium the scolex is set off early from the the rest of the body (Figs. 
57, 70) and soon afterward is completely invaginated. The Proteoceph- 
alus procercoid bears a close resemblance to the plerocercoid found in 
the intestine of the final host. This is not true of the procercoids of Cor- 
allobothrium, in which a decided transformation takes place in the pro- 
cercoid before the plerocercoid stage is attained (Figs. 58, 59, 68). In 
every case the Proteocephalus procercoid has a greater length than that 
of the Corallobothrium procercoid. In the three species of Proteocephalus 
whose life-cycles have been studied, only one intermediate host is re- 
quired. The evidence is fairly substantial that at least one species of Cor- 
allobothrium, C. fimbriatum, may use two intermediate hosts for its 
development. The striking differences in the development of these ces- 
todes, combined with the differences in adult morphology, should be, in 


307] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 53 


my opinion, sufficient evidence to satisfy the most critical worker that 
Corallobothrium should remain a separate genus. 


COMPARISON WITH DIPHYLLOBOTHRIUM LATUM 


The species of Proteocephalus have been compared with those of 
Corallobothrium from the point of view of their development. A similar 
comparison between Diphyllobothrium latum and the species of Corallo- 
bothrium is of interest. As described by Janicki and Rosen, D. latum 
develops in the following manner: The ciliated larva is ingested by 
Cyclops strenuus. Within a few hours the oncosphere has gained the body- 
cavity of the Cyclops. The larva remains attached by its hooks to the wall 
of the intestine 10 to 15 days, during which time it loses its contractility. 
It becomes elongated and at the end of 6 to 8 days development measures 
0.10 to 0.15 mm. At the end of from 8 to 12 days the larva shows cal- 
careous bodies, longitudinal and transverse musculature and a strongly 
developed cuticular covering. On the twelfth day after beginning de- 
velopment the opposite poles of the body show differentiation. By 
the twelfth to fifteenth day a special bladder appendage with the em- 
bryonic hooks upon it is formed. Now the larva measures from 0.35 to 
0.40 mm. At the end of 15 to 20 days the larva has attained a length of 
from 0.5 to 0.6 mm. Then the bladder gradually degenerates and dis- 
appears. At the time of the formation of the bladder an evagination 
occurs at the opposite end. Later this structure is capable of being in- 
vaginated. At the end of development in the Cyclops the larva is covered, 
especially its anterior portion, with bristle-like processes. At this stage 
the body is highly contractile. The scolex and excretory vesicle are not 
formed by the larva while in the Cyclops. These two organs make their 
appearance in the second intermediate host. 

The behavior of the oncosphere of Corallobothrium in some particulars 
is decidedly different from that of D. latuwm. In the former the oncosphere 
does not remain attached to the intestine of the Cyclops by its hooks. 
The hooks may be seen in action at any time after the oncosphere has 
gained the body-cavity. Neither does the larva lose its contractility. 
The development of Corallobothrium larvae is much more rapid, as they 
attain their greatest length at the end of from 6 to 8 days. In D. latum 
the bladder bears the embryonic hooks; but in Corallobothrium the bladder 
may or may not possess the hooks. The scolex and excretory vesicle 
are not formed in D. Jatuwm until the larva has developed in the second 
intermediate host. In Corallobothrium the scolex is formed early, by 
the tenth or twelfth day. The species of the last-named genus do not pos- 
sess the covering of bristle-like processess noted for D. latum. The Corallo- 
bothrium procercoid is never more than two-thirds the length of the 
procercoid of D. Jatum. Another interesting difference is found in the 


54 ILLINOIS BIOLOGICAL MONOGRAPHS (308 


appearance of the excretory vesicle. In D. latum this organ does not 
occur until the plerocercoid stage, but in Corallobothrium giganteum it 
arises during the development in the Cyclops. It was not observed in 
Corallobothrium fimbriatum until the plerocercoid stage. This might be 
expected since the procercoid development of C. giganteum covering 10 to 
12 days is more rapid than that of C. fimbriatum which takes 14 to 15 days. 

Janicki and Rosen never found more than one mature procercoid in 
the same Cyclops. I have observed as many as three mature procercoids 
and three individuals representing 10 days development in the same host. 
The largest number of larvae reported for D. Jatum in a single Cyclops 
was from 8 to 10. As many as 18 were found in a single host infected with 
the eggs of Corallobothrium fimbriatum. 

This comparative study of the Bothriocephalid, Proteocephalus, and 
Corallobothrium procercoid furnishes evidence indicating an intermediate 
postition for Corallobothrium between the Bothriocephalids and _ the 
genus Proteocephalus. First, in the Bothriocephalids the cercomer is 
always present and nearly always bears the embryonic hooks, but in 
Proteocephalus it may be wanting entirely. When present, however, it 
never bears the embryonic hooks. In Corallobothrium the cercomer is 
always present and it may or may not possess the hooks. Second, the 
procercoid of the Bothriocephalids bears little resemblance to the plero- 
cercoid, whereas the Proteocephalus procercoid is almost identical to the 
plerocercoid while the procercoid of Corallobothrium does not bear a close 
resemblance to the plerocercoid stage. The Bothriocephalids usually 
require two intermediate hosts, the Proteocephalids only one, while 
Corallobothrium may or may not use two intermediate hosts. 


EARLY DEVELOPMENT IN BOTHRIOCEPHALIDS AND PROTEOCEPHALIDS 


Observations on the early development of the Proteocephalids are in 
close agreement with those on Diphyllobothrium latum, Triaenophorus 
nodulosus, Abothrium infundibuliforme and Ligula simplicissima. There 
are, however, a number of differences that should be considered. The 
egg of the Proteocephalids when extruded from the uterus, possesses a 
gelatinous outer covering which enables it to float about for a time before 
sinking to the bottom of the stream or lake. Within this gelatinous material 
is found a second membrane (shell) which contains a quantity of granular 
material and the fully-formed oncosphere which may or may not be in- 
vested with a third membrane. The oncosphere of the Proteocephalids 
is held a prisoner within its membranes until liberated through the agency 
of the first intermediate host. The egg of each of the Bothriocephalids 
mentioned above, closely resembles that of the trematodes. A gelatinous 
covering is absent. The oncosphere may be fully developed before the egg 


309] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 55 


is extruded from the uterus, as in Abothrium infundibuliforme; or an 
incubation period of varying length may be necessary after the egg is 
extruded, as in Diphyllobothrium latum. When the egg is mature it is 
composed of an outer shell, which I consider to be homologous with the 
second membrane of the Proteocephalid egg, and a layer of cells which 
envelops the oncosphere. This layer of cells is probably homologous 
with the innermost membrane reported in most Proteocephalid eggs. 
Contrary to what occurs in the Proteocephalids, the egg of the Bothrio- 
cephalid hatches, thus liberating the oncosphere. The layer of cells envelop- 
ing the oncosphere may be provided with cilia, as in D. datum, or the cilia 
may be wanting, as in A. infundibuliforme. 

A comparison of the size of the oncospheres of certain Bothriocephalids 
and Proteocephalids is given in the following parallel columns. 


Bothriocephalids Sizein Authority Proteocephalids Sizein Authority 
micra micra 
Diphyllobothrium latum 22to30 8 Janickiand Proteocephalus 
Rosen filicollis 27(estd) Meggitt 
Triaenophorus nodulosus 22to24 Rosen P. torulosus 20 to30 Wagner 
Abothrium infundibuliforme 50to60 Rosen P. percae 28 to35 Kuczkowski 


Corallobothrium 
fimbriatum 20 Essex 
C. giganteum 13to16 86“ 


It is of interest to note that the first intermediate hosts of the Pro- 
teocephalids and Bothriocephalids considered in this paper are found 
exclusively among the copepods. The species recorded in the following 
columns have been studied experimentally. 


PROTEOCEPHALIDS 


Parasite 


Proteocephalus filicollis 


P. torulosus 


P. percae 


P. longicollis 


Corallobothrium fimbriatum 


C. giganteum 


First Intermediate Authority 


Host 
Cyclops varius Meggitt 
Diaptomus castor 


Cyclops strenuus Wagner 


Cyclops strenuus 
C. serrulatus 


C. oithonoides Kuczkowski 


Cyclops strenuus 


C. serrulatus Kuczkowski 


Cyclops bicuspidatus 
C. serrulatus 


C. prasinus Essex 


Cyclops serrulatus 


C. prasinus Essex 


56 ILLINOIS BIOLOGICAL MONOGRAPHS 


[310 


BoTHRIOCEPHALIDS 


Parasite 
Diphyllobothrium latum 


Diphyllobothrium latum 
Triaenophorus nodulosus 


Abothrium infundibuliforme 


Ligula simplicissima 


First Intermediate 


Host Authority 
Cyclops strenuus 
Diaptomus gracilis Janicki and 

Rosen 

Diaptomus oregonensis Essex 
Cyclops strenuus 
C. fimbriatus Rosen 
Cyclops strenuus 
C. serrulatus Rosen 
Cyclops strenwus 
Diaptomus gracilis Rosen 


There may be some significance attached to the fact that Cyclops 
strenuus acts as the first intermediate host of all the Bothriocephalids 
and three of the Proteocephalids; that Cyclops serrulatus is found among 
the first intermediate hosts of both groups. 


311] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 57 


AFFINITIES OF THE PROTEOCEPHALIDS 


A study of the life-cycle of organisms has long been used as a guide 
to their true affinities. Adult morphology alone is not always a reliable 
criterion of true relationship. Certain forms, the barnacles and ostracods 
for example, which were thought to be widely separated phylogenetically 
on the basis of the adult structure, have been brought close together 
through the discovery of their developmental cycles. The lack of com- 
plete knowledge on the life-cycles of the vast majority of the Cestoda 
has necessitated a grouping based almost entirely on adult morphology. 
In recent years sufficient data have been gathered on the development of 
the fish cestodes to justify pointing out certain apparent affinities. 

By the discovery of the life-history of Diphyllobothrium latum Janicki 
and Rosen (1917) were able to indicate more fully the relationship between 
the Digenea and the Bothriocephalids which was first suggested by 
Leuckart (1886). This is shown by the following points: 


1. Similarity of the eggs 

2. Existence of a uterine pore 

3. Structure of the larvae 

4. Two intermediate hosts 

5. Resemblance between the miracidium and coracidium 
6. Resemblance between the cercaria and the procercoid 


On the basis of the points just enumerated, Diphyllobothrium latum, 
Triaenophorus nodulosus and Ligula simplicissima show a clear affinity 
to the Digenea. The development of Abothrium infundibuliforme as 
described by Rosen (1918) differs from the three Bothriocephalids just 
mentioned in respect to the fifth point. Although the egg of A. in- 
fundibuliforme goes through the process of hatching, it does not give 
rise to a ciliated larva or coracidium. In this respect it is further removed 
from the Digenea than the other Bothriocephalids whose life-cycles are 
known. 

Using the points enumerated above as criteria of the relationship 
between Bothriocephalids and Proteocephalids, the first variation from 
the usual Bothriocephalid condition if found in Abothrium infundibuli- 
forme; the eggs of which show an intermediate position. Since the eggs 
of the last-named species contain a fully formed oncosphere when extruded 
from the uterus, they resemble the eggs of the Proteocephalids; but 
since the eggs of A. infundibuliforme give rise to unciliated larvae, they 
differ from the usual Bothriocephalid condition. Except for the first and 


58 ILLINOIS BIOLOGICAL MONOGRAPHS {312 


Afth points there is substantial agreement between Corallobothrium jim- 
briatum and the Bothriocephalids. Proteocephalus percae agrees on the 
second, third, and sixth points, but P. filicollis and P. torulosus differ in 
all points except the second and third. Therefore, starting with a typical 
Bothriocephalid such as Diphyliobothrium latum, there is a striking series 
of transitions in arriving at the typical Proteocephalid condition. 

The first variation is found in Abothrium infundibuliforme; whose 
larva is unciliated. Next comes Corallobothrium fimbriatum, which varies 
from the former only in the type of egg. Then Proteocephalus percae 
follows, varying from C. fimbriatum in the rather rudimentary condition 
of the cercomer. Finally, in Proteocepalus filicollis and P. torulosus a condi- 
tion is reached which differs from P. percae in that no cercomer is re- 
ported and thereby the resemblance of its procercoid to the cercaria is 
lost. 

On the basis of the scolex, Fritsch (1886) suggested that Corallo- 
bothrium solidum represented a connecting link between the Bothrio- 
cephalids and the Taenias. In the light of the knowledge of the develop- 
ment of the two groups there is much to support his suggestion. La Rue 
(1914), after a thorough study of the species of the Proteocephalidae, 
found them closely related morphologically to the Tetraphyllidea. How- 
ever, in their development the Proteocephalids show a close affinity to 
the Pseudophyllidea. 


SUMMARY 


1. An intensive study has been made of two new species of Corallo- 
bothrium. Both species are parasitic in /ctalurus punctatus, Ameturus 
melas, and Leptops olivaris. Woodland’s proposal (1925) to delete the 
genus Corallobothrium has not been accepted. The genus has been re- 
tained and the two new species have been designated as Corallobothrium 
giganteum and C. fimbriatum. A description is given of the adult anatomy 
of each parasite. The scolex of C. giganteum is subject to wide variation. 
Amphitypy occurs in the arrangement of the interovarial organs of both 
species. A new role has been suggested for the excretory system. 

2. Particular attention has been paid to the degree of infection and 
seasonal occurrence of these cestodes in Ictalurus punctatus. Among 
130 adults of this species nearly 70 per cent harbored one or the other of 
these cestodes or both. The adult parasites occurred only from spring 
to fall but the plerocercoid stage was present in the final host through- 
out the year. 

3. The life-cycle of each cestode was studied experimentally. In- 
fection was produced in Cyclops serrulatus and C. prasinus by feeding 
eggs of Corallobothrium giganteum. Positive results were obtained by 
feeding the eggs of C. fimbriatum to Cyclops bicuspidatus, C. serrulatus 


313] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 59 


and C. prasinus. The complete development of both parasites in the first 
intermediate host has been described. The procercoid of Corallobothrium 
giganteum reached maturity in the Cyclops by the twelfth day after the 
Cyclops had been exposed to the eggs. The development of the pro- 
cercoid of Corallobothrium fimbriatum required from 12 to 14 days. 

4. Observations on the feeding habits of Cyclops reveal that organic 
debris and cercaria are eaten and that protozoa are consumed in large 
numbers. 

5. Cyclops infected with mature procercoids of Corallobothrium 
fimbriatum were fed to minnows (WNotropis blennius). Larvae were re- 
covered from the body-cavity of the minnows 3 days after feeding. Sec- 
tions of an infected minnow showed the presence of the larvae in the 
intestine, within the coelom, and in the musculature. 

6. Ictalurus punctatus measuring from 2 to 3 inches in length were 
found to harbor adult Corallobothrium fimbriatum. 

7. Cyclops infected with the procercoids of Corallobothrium fim- 
briatum were fed to minnows. The minnows were fed to Ameiurus melas 
and the larvae of Corallobothrium fimbriatum were recovered from the 
intestine of A. melas. 

8. The evidence indicates that catfish may be infected with Corallo- 
bothrium fimbriatum either by ingesting infected Cyclops or by feeding 
on infected minnows. 

9. A comparison of Bothriocephalid and Proteocephalid develop- 
ment indicates a close relationship between the two groups. 


60 ILLINOIS BIOLOGICAL MONOGRAPHS (314 


BIBLIOGRAPHY 


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Kraemer, A. 
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Linton, E. 
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317] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 63 


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ZSCHOKKE, F, 
1884. Recherches sur l’organisation et la distribution zoologique des vers parasites des 
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1903. Die Arktischen Cestoden. Fauna Arctica, 3: 1-32; 2 pls. 


are used: 


cip 
dj 
def 
exd 
exv 
fs 
ml 
mr 
nl 
nr 
od 
oot 
ov 


ILLINOIS BIOLOGICAL MONOGRAPHS 


[318 


EXPLANATION OF PLATES 


All figures, except 39 and 41, were drawn with the aid of the camera lucida. The value 
of the scale projected is indicated in the explanation of each plate. The following abbreviations 


cirrus-pouch 

ductus ejaculatorius 
vas deferens 

excretory vessel, dorsal 
excretory vessel, ventral 
foramina secondaria 
longitudinal muscles 
muscle rhomboid 
lateral nerve 

nerve ring 

oviduct 

ootype 

ovary 


00c 
rs 
sm 
tt 

ut 
ull 
utp 
utop 
va 
val 
vt 
vid 
vide 
vir 


oocapt 

receptaculum seminis 
sphincter muscle 
testes 

uterus 

lateral uterine pouches 
uterine passage 
ventral uterine pore 
vagina 

lower vagina 

vitellaria 

vitelline ducts 
vitelline duct, common 
vitelline reservoir 


319] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 65 


PLATE I 


66 


_ILLINOIS BIOLOGICAL MONOGRAPHS 


EXPLANATION OF PLATE I 


The value of the scale projected on each figure equals 0.3 mm. 


Fic. 1. Corallobothrium fimbriatum, scolex of adult, toto. 
Fic. 2. C. giganteum, apical view of scolex. 


Fic. 3. C. fimbriatum, organs of interovarial space, toto-mount. 


Fic. 
Fic. 
Fic. 
Fic. 
Fic. 
Fic. 
Fic. 
FIG. 
Fic. 
Fic. 
Fic. 
Fic. 


shown, 


C. fimbriatum, scolex of adult, toto, more expanded than fig. 1. 

C. giganteum scolex, showing apical prominence. 

C. giganteum, fully extended proglottid from near posterior end. 

C. gigantewm, protruded cirrus, cirrus-pouch, vagina and vas deferens, toto. 
C. fimbriatum, ovary of immature proglottid, toto. 

C. fimbriatum, ovary of mature proglottid in same chain as fig. 8. 


. C. giganteum, dorsal view of the scolex, much contracted, toto. 

. C. giganteum, apical view of the same scolex as in fig. 10. 

. C. fimbriatum, ovary of immature proglottid, toto. 

. C. fimbriatum, ovary of mature proglottid in same chain as fig. 12. 
. C. fimbriatum, frontal section of immature segment. 

. C. giganteum, expanded scolex, toto. 


[320 


Vitelline ducts are not 


ILLINOIS BIOLOGICAL MONOGRAPHS VOLUME XI 


15 


ESSEX CORALLOBOTHRIUM PLATE I 


321] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 67 


PAE it 


68 


ILLINOIS BIOLOGICAL MONOGRAPHS (322 


EXPLANATION OF PLATE IT 


The value of the scale projected on each figure equals 0.3 mm. except 
fig. 24, on which it equals 0.1 mm. 


Fic. 16. Corallobothrium giganteum, frontal section of contracted scolex showing excretory 
vessels much reduced in apex. 


Fic. 
Fic. 


Fic. 
Fic. 
Fic. 


Fic. 
Fic. 
FIc. 
Fic. 
Fic. 
Fig. 
Fic. 


uli 
18. 


C. giganteum, cirrus fully protruded, drawn from living specimen. 
C. gigantewm, cross-section near apices of suckers, showing sphincter about suckers 


and muscle fibers connecting each pair of suckers. 


19. 
20. 
21. 


C. fimbriatum, sagittal section of much contracted scolex. 
C. giganteum, cross-section at level of cirrus-pouch. 
C. giganteum, frontal section of expanded scolex showing distended excretory vessel 


in apex. 


22. 
23. 
24, 
20 
26. 
27. 
28. 


star. 


C. fimbriatum, frontal section of expanded scolex. 

C. giganteum, fully expanded ovary and associated organs, toto. 

C. giganteum, procercoid showing scolex partially protruded. 

C. fimbriatum, ripe proglottid, toto. 

C. giganteum, mature proglottid. 

C. fimbriatum, cross-section. 

C. giganteum, cross-section near posterior limit of suckers showing part of muscle 


ILLINOIS BIOLOGICAL MONOGRAPHS VOLUME XI 


De th, 
US 


XY, 
bit 


ESSEX CORALLOBOTHRIUM PLATE II 


323] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 69 


PLATE III 


70 ILLINOIS BIOLOGICAL MONOGRAPHS (324 


EXPLANATION OF PLATE III 


The value of the scale projected on each figure equals 0.3 mm. except on 
figs. 39, 40 and 41, on which it equals 0.03 mm. 
Fic. 29. Corallobothrium fimbriatum, frontal section of mature proglottid. 
Fic. 30. C. gigantewm, frontal section of scolex contracted as in fig. 10. 
Fic. 31. C. fimbriatum, cross-section showing extent of uterus. 
Fic. 32. C. fimbriatum, frontal section of dorsal region of mature proglottid. 
Fic. 33. C. giganteum, frontal section of an expanded scolex showing the sphincter muscle 
of the suckers as a knob-like structure. 
Fic. 34. C. giganteum, drawing made from three frontal sections. The proglottids shown 
here are three segments posterior to those shown in fig. 38. 
Fic. 35. C. gigantewm, cross-section through scolex near apices of suckers. 
Fic. 36. C. fimbriatum, frontal section of vagina and protruded cirrus and cirrus-pouch. 
Fic. 37. C. giganteum, frontal section of contracted scolex, sphincter of sucker shaded. 
Fic. 38. C. gigantewm, drawn from three frontal sections. The excretory vessels are much 
less distended than those shown in fig. 34. 
Fic. 39. C. giganteum, reconstruction of interovarial organs. 
Fic. 40. C. giganteum, cross-section of dorsal excretory vessel. 
Fic. 41. C. fimbriatunt, reconstruction of interovarial organs. 


ILLINOIS BIOLOGICAL MONOGRAPHS VOLUME XI 


ESSEX CORALLOBOTHRIUM PLATE III 


325] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX 71 


PLATE IV 


72 ILLINOIS BIOLOGICAL MONOGRAPHS {326 


EXPLANATION OF PLATE IV 


The value of the scale projected on each figure equals 0.03 mm. except on 
figs. 47 and 51, on which it equals 0.3 mm. 
Fic. 42. Corallobothrium fimbriatum, mature egg. The funnel-like cavities at each pole are 
indicated. 
Fics. 43 and 46. C. fimbriatwm, mature eggs in which the middle membrane is ruptured. 
Fics. 44 and 45. C. fimbriatum, oncosphere 10 minutes after being removed from the body- 
cavity of the Cyclops. 
Fic. 47. Cyclops bicuspidatus, containing 8 C. fimbriatum larvae 3 days after feeding of eggs. 
Fic. 48. C. fimbriatum, larva removed from Cyclops 3 days after feeding of -eggs. 
Fics. 49 and 50. C. giganteum, mature eggs. 
Fic. 51. Cyclops bicuspidatus, which contained 18 C. fimbriatum larvae 6 days after feeding 
of eggs. 
Fic. 52. C. gigantewm, tracing of oncosphere seen in the body-cavity of Cyclops 8 hours after 
feeding of eggs. 
Fic. 53. C. fimbriatum, larva seen in abdomen of Cyclops 24 hours after feeding of eggs. 
Fic. 54. C. fimbriatum, 4-day old larva. 
Fic. 55. C. fimbriatum, 36-hour larva as seen through body wall of Cyclops. 
Fic. 56. C. fimbriatum, larva about 10 days old. 
Fic. 57. C. fimbriatum, larva from 10-11 days old. 


VOLUME XI 


ILLINOIS BIOLOGICAL MONOGRAPHS 


CORALLOBOTHRIUM PLATE IV 


ESSEX 


327] STRUCTURE AND DEVELOPMENT OF CORALLOBOTHRIUM—ESSEX = 73 


PLATE V 


74 ILLINOIS BIOLOGICAL MONOGRAPHS [328 


EXPLANATION OF PLATE V 


The value of the scale projected on each figure equals 0.05 mm. except on 
figs. 62, 66, 67 and 69, on which it equals 0.2 mm. 
Fic. 58. Corallobothrium fimbriatum, procercoid 12-13 days old with the bladder appendage 
still attached. 
Fic. 59. Plerocercoid taken from intestine of 7. punctatus. 
Fic. 60. C. fimbriatum, larva somewhat contracted taken from body-cavity of Notropis 
blennius to which infected Cyclops had been fed. 
Fics. 61 and 63. C. gigantewm, forms found in Cyclops which also contained procercoids as 
shown in fig. 68. 
Fic. 62. C. fimbriatum, very young plerocercoid. 
Fic. 64a, b, c, etc. C. gigantewm, procercoid, outlines indicate shapes assumed before evagina- 
tion of scolex. 
Fic. 65. C. fimbriatum, cross-section of NV. blennius intestine with larva outside of intestinal 
wall. 
Fic. 66. C. giganteum, procercoid with scolex evaginated, from preserved material. 
Fic. 67. Cyclops serrulatus, containing 3 procercoids of C. giganteum. 
Fic. 68. C. gigantewm, mature procercoid. 
Fic. 69. C. fimbriatum, a much enlarged drawing of larva shown in fig. 65. 
Fic. 70. C. giganteum, larva from 8-10 days old. 


ILLINOIS BIOLOGICAL MONOGRAPHS VOLUME XI 


CORALLOBOTHRIUM PLATE V 


Anat 
Pie helapmmeneoetannnst 


rou ane 
Tyletiey ava: 


Cetera) 


bap nie nes 
PPNARLCECTIIOR AT Ltt ete 
Peron cite ey: 


itr} HUAN asbepeath 
CHAM a nhite Dee 
Praha tr arenes 


na ond We Bah ky ee UNIVERSITY OF ILLINOIS- URBANA 


3 01 12 0650971 04 


baa 
iano ie 


Leet i 
bp 


Ap bigsd nr pat Poa ; 
pe ere i et HEP sear hetet mies) CE 
A 1 taehart gabap baht ait ntarrsat ont Peli wet hernia : 
bbe Wu mh Pht Ah nomtnerymrmyDeL OH ON rey tee titi Heirons hin ir rahe AD * 
enue tne getat pany I atcLaba pent geuomyeel telah : 
seve menace pcaonet postion vite zy 
Veet OA taseaneg ane Hany edad hehe ; 
ebb ontne psy ye iy ) Int enh an alk ET peonsnce hi H a tian 
, er iirishty eee ya Reeaeeuse i } poets 
Wp bee fi tabby neat ye Pee racer el tad ae a 
Haba iene ey a i f / 
IFiehee be Pro PAMNYI6 ee ‘ 
dire en erisrn ai i vie 
opment isin Mg w Phyel 
ni ee eater A ; 
Tabs haere Went 
Ry aaah ear ke ata ike 
i Haein : 
ane MU EEMivur oui iappepalite sil HOt i i 
vier Mee Ther { rein 
M badd Linde Me in bovibebapbE LAO MRI IRDAR I Ais ory 
ine wey yeaUnindawey ed at a neta rt : ates 
Mibhnedee ‘henheriapeane ab soeyeNsiley pb mn yt ue { 
seo re seoee Her ebHOULO KE # hee é ¢ i 
vied tN crwee’ infer the Heal, fh eat 
brig i xc eiinad als atta 
Sida ge cty ret ts anda the i omibie ’ 
on i Prokaj ine] ee porate jed Jopreptendy ! 
dpe meee UE Us LSaen fanaa a Chg ecin ty iu Meshcrste 7 
rpeateey: jefewar abi De bythe Fey 1 ert hs ty His yh PAE TK 
eoprbaya Drab aD age DH “i Tin oe op tan Dinabey - . 
sscae pede pratt ne Seb} . whekes t 
miei eee ya Nainete ltr pial ‘ 
teetityiob eas Hit detest met taatt Lage 
Lidaticiatcetilabepneaieaatieswegt ttn db meat ‘ Ape : wet tt Tova ks 
{rots signa ms sont tity en) tie mr ttetyy EA TWebwn (i yeh ke coer hat 
EI ta at vA siti nae ges i letid bik hy ds Sieshacsspstniueneer ge pists 
Sates gs en ei) tently e } 
Thee iwecatmi/ deat Hf HA HSER naanaTA i aver we hd ctor Fouig : “ 
itinsonen Binulaentie at ainehd Teen ecedety MV Riiibehapele thy Vat bunt WpAiW feb aba delipheiom. tus : 
Lapel PANDA cantante esas heer aiaiaybp Hii yeaenvn raya ory aaaeDant ane) ASAD thoes bs hepibouy . : 
AN Tunatboas(stan'eace rn tiseaiteit atsere atria SUCH tEM ap sliveniret bu 14 
ITE AHN ET ee RE tase is * : 
He itisctisab aca Nagel fi, 2 1 
Pabruipbant pati, dh eynnie ra be i an 
(ut : 
er ete th Ne * * 
Denehsties MY ey Hetpedene } yee 
sere rivanirtakcatee Teele ies ; f 
Hold ebraMeerietig wt int GE i 
Mu es frie e 7 
ENB d poe ara abe tonsa os 
sins pith f ithe : stn webshots 
Sibi atte ( rie he 
f ; i x : 
rile cs : \ 
ets bey ars - ‘ " Hier : 
ae en A subtree matcwur weil - 2 
Feanigeatnieg heahertD qreairaaiyene : ¥ : 
Pith enmiharts tOaP betty } 


ietnebrit ae Peas hope psi iaé 


sovindarnngmoes Utes mht at 
are 


oath Deb Lat 
dohabe tat bsidwy| 


wregaae 


Hele nid | Daehn Bh me eqns viyey nutolia cy Y t ei rf 

here UML N RY EH HDL \ HS aieuurar iasrapeyraec rit | mats ht THUY SMe eile iedetHieget 

waged ont caneegnte tg he Satire he heh MIURA hat ba aa eat NaN i A 
retina wld aah eele ta Hi aertbe aed rate RucuptbnE ALL wah hin 

Saud att Hauer sts ya dena Sean ao DU abl Ma 9A WE Vay oth eon tan 


re barat eel nuaturest et 
seep syst Vina pe ay 
la Seepage bay te 


i tiynay jest 9 
repeated ny np hpmetCaite te eibebt ss 
Wee UI OHL AIRED EH Meaty EA 
nose aa 

elo eaten 


Ah LUSH oR Nw OS bi WORE OvaoMoba gel Mote 


Geren To aa eee a 
yy" Hititatad rian: prastiupmiiRen 


Hemten Ansel 


elit 

Hibinacee are 
SEM NS a 
perileh ae a) seurshnnlt verti 
Cite idtok 


in tate 
Sule writhve 


arta hn acuirer aay Sweeney het 


vie tA 
, vit SReTS olan 
* 


‘aie 
Matynss hii 


Srteter bye yates heen 
Maher rail ras Rati Spann a ei 
spats eer i alae nabatt 
tye Nitti cures besten? : vets 
i mimetic Witanht teenie taleaseni pine 
itaeueae TMVAbaLH AbAyani ood DeubaretaeT ee Roorles ta macasone ttl 
et Meaiilins sti cry tabdegit TRAN lity tt ea ye A 


ON bh od lt ob 
my eee bits 


i 

4) heave ge) vil! 
ot vei) era TO H 
ingqvis stat? “ ye 


<a n nein Larat area ie mid 
a yeVorirasean tals maicoues it sienrentt sc titan banal ertcaet touhany 
span iesce ai it taiiNisu alareraek 5 deeb sheila rete de ie hy 


ecapepetty wlgnnomrpearer bvanteye 
inreaeh fer eapat cata) vita a 
Tolan iat Sat 


ees tabi 


Vente 


ihren. * Asante 
TWH peje At hg 
teerueararnsiiel 


ead ere at 


bade neat hel trae day Sivan tv 
Maren erratitactatte Deut patton eer Tiddt oh ep thegetiel ols apelin 


we 
ft perks) Gry Ti Ferbhuvieyy gahee 
aenatbes Maroon serie 


sal pn #1} he slab iis aeanes toe true annoy 


tise 


ibe ant aap ite eee torboh iy niet etd Ne tat te 
Ce td srt aiisiier th hire iain bs eenoie oe Muu ptnrielectayet 
aicemisa nt sat iG tans SRM eat vit! Peart mw aco att oabatrt uefa pw aalcn een ita 
AP ri mnitheticden penoieen tice “ auitibeeed Heacuablond Invi Bineue ed nett shite spose is 
ten nd wit itisehe vibceiel ish wien Nel cpsinbee ileal 


Woon 


i} eet DyPe tropes Hisel obeiewc ntl aT 


ines uerinae at 


Bo hiene 


nite 
Ni abespeee aM atiab ns eats ch 


FHC RC Gish elie ticten nnd 
reek at eben ete a Herbed ing 


Mah UCMEebel Werke yelieeeel bra yereee 
Sieh 


stiMa inh toepebearianepredte 
oh Fs thee Y 


DeSean 
ity eaten 
tt ens ene ita ii eit 


eae 


MbOPHOLGANIDNE D2 IPT 


Wrbiaeests 


a £ 
teh aa AGT UE at Guba — iin Bnet 
sett PA shine ivisbavienyes tis ie NGS toatl OCCA Ret ie He Leppegiey sti 
f 


Maesbeatays Denpitbtey UAL whist te) 
TS rd { UG 7 \ 


maine i ai 


rn 
i 
ae 


Hires orwisb feb we di iat: 
ste 


pnfrirb bret 
Nae erand sbreet 
Pre alten ed 
Wi jive be veetgee bbw 


rae 

2 Fake Hote id a 
So i 
eatieaoes 5 


7 yore! of 
aft 


toch heer rs batt 
erty Watt rtd aie 


ahs ie jean evades Ube b naet as asians 
ata Binteia ronnie igri Caayvegeeaguan Nnesoefvenptya fv 
i ee aaeaets at arluistetnst f 


oad . ‘} CAND he aal 
( ee ” ee Ys yt ANE ee 
gitar fatesgic gt ogni tayceuiptee oe ieateemteuaten 


rane feab eth iy . 
auibevtiecraues meiahe yet pemley Hrd Mrrosiiaae iteeeanonta hvac 
at Meat Pia sefeangh rh Tred veareyitetie ht Hira ot 


het ain 


“vrtewiae 


ape brah tay 


af iy ae Feit stia seb ae uae eh seca stabs Me inner atoene on Esp cli heya sa 
idea aia aaah Pea can sea ‘ine ne esti Obs 
pay idpehicngen enh sit Neenah BELT haw 
Minhueniegtaaiatiesaua tte yenecy nitrate Fanti 
at Nagel sean Nels Hit keer 
i Hens ae atu tiecprisapifbeesce 
Cag a gt Bekayent beeen eieaned she 


WA adeecahsn opal te OBIT Ne Peitkl ie deLieiye nel 
FeLi AE 
wis 


Sa eonununrey id 


iy 


seal eapeaeg 
Seve avitegeaeh 


is sa neti cece eye 
{i ative sh ephatres ate fee rie rata oct 
Sibert a tireutes Mbetbanaiel toss debeitanen 
tiga i tar ibhen Uaarteets heket tar 
coat roster tes Lah tiadade ete 
sores yl 4 i a Nid reners hur oribkt eon 
ti tei mast ; pavbiousasicamerertaetertts 
i in pb aaeny asian niece etait ur hy 


i Hacer tan 


Hy BF Hea ALR Eni eb A Re weRl Wty oT 


a iteeksvahige ve ashe sel ttt gebed main sides bear efile 
i salvonchutpaes iat i Sie dee cal 
Ringupeirn ete Adina ihe vorke arb Uebey 
ie tea ashen eee Hi oraeb thE ihe dets 
isilinltateh i esien at ieae ld eshantsesrelbpeaetcne (Si 


: 
UNO TRUSTE Dae cence epaetn a MLR Maus bei 
NT RTA Sach sosheibe capi eurnnecasbee bi vlieUnclie isa tunnaee) i 


